Registration Dossier
Registration Dossier
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EC number: 240-539-0 | CAS number: 16484-77-8
Adsorption / desorption
Administrative data
Link to relevant study record(s)
- Endpoint:
- adsorption / desorption: screening
- Type of information:
- experimental study
- Adequacy of study:
- key study
- Study period:
- 19 January 2010 to 21 June 2010
- 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)
- Version / remarks:
- 2000
- Deviations:
- no
- Qualifier:
- according to guideline
- Guideline:
- other: EPA OPPTS 835.1230
- Deviations:
- no
- GLP compliance:
- yes (incl. QA statement)
- Type of method:
- batch equilibrium method
- Media:
- soil
- Radiolabelling:
- yes
- Analytical monitoring:
- yes
- Details on sampling:
- - Concentrations: All concentrations
- Sample storage before analysis: Samples were stored refrigerated until analysis. All adsorption supernatants analysed during the preliminary experiments were profiled by HPLC within 3 days of their generation. - Matrix no.:
- #1
- Matrix type:
- sandy loam
- % Clay:
- 13
- % Silt:
- 18
- % Sand:
- 69
- % Org. carbon:
- 3.13
- pH:
- 5.8
- CEC:
- 12.1 meq/100 g soil d.w.
- Bulk density (g/cm³):
- 1.12
- Matrix no.:
- #2
- Matrix type:
- clay loam
- % Clay:
- 33
- % Silt:
- 26
- % Sand:
- 41
- % Org. carbon:
- 3.71
- pH:
- 7.3
- CEC:
- 25.6 meq/100 g soil d.w.
- Bulk density (g/cm³):
- 1.13
- Matrix no.:
- #3
- Matrix type:
- sandy clay loam
- % Clay:
- 31
- % Silt:
- 22
- % Sand:
- 47
- % Org. carbon:
- 3.07
- pH:
- 5.7
- CEC:
- 22.9 meq/100 g soil d.w.
- Bulk density (g/cm³):
- 1.12
- Matrix no.:
- #4
- Matrix type:
- loamy sand
- % Clay:
- 5
- % Silt:
- 8
- % Sand:
- 87
- % Org. carbon:
- 2.9
- pH:
- 5.7
- CEC:
- 10 meq/100 g soil d.w.
- Bulk density (g/cm³):
- 1.36
- Details on matrix:
- JUSTIFICATION OF SOILS
Four agricultural soils collected from various sites in the UK and Germany were used for the study. The soils were selected to cover a range of organic matter content, clay content and pH.
The soils used in this study are listed below:
- Calke sandy loam
- South Witham clay loam
- Lockington sandy clay loam
- Hagen (Refesol 04-A) loamy sand
All soils were classified under USDA system. The soils were certificated by Agvise Laboratories, Northwood, North Dakota, USA.
As degradation of [14C]-test material was observed in the soil: solution ratio investigations, the remaining preliminary determinations (adsorption and desorption equilibrium) and the definitive test were conducted using sterilised soils. Soils were sterilised (Isotron plc, Swindon, Wiltshire, UK) by exposure to gamma irradiation at a minimum dose of 25 kGy.
COLLECTION AND STORAGE
- Geographic location: Calke sandy loam collected from Derbyshire, UK on the 5th January 2010, South Witham clay loam collected from South Witham Quarry, Lincolnshire, UK on the 5th January 2010, Lockington sandy clay loam collected from Lockington Grounds Farm, Leicestershire, UK on the 5th January 2010 and Hagen (Refesol 04-A) loamy sand collected from Schmallenberg, Nothrhine-Westphalia, Germany on the 5th January 2010.
- Collection procedures: Calke sandy loam: By spade after removal of surface turf layer; South Witham clay loam: By spade form shelf cut into top soil; Lockington sandy clay loam: By spade form shelf cut into top soil; Hagen (Refesol 04-A) loamy sand: By spade after removal of snow.
- Storage conditions: All four soils were stored at ca. 5 °C in a plastic bag in the dark.
- Soil preparation: The soils were dried slightly if needed, before being passed through a 2 mm sieve and fully characterised, with respect to particle size distribution, cation exchange capacity/exchangeable cations, organic matter content, pH, bulk density and water holding capacity. On arrival at the testing facility, all soils were stored in accordance with ISO/DISS 10381 Part 6 to maintain viability.
The moisture content of each soil was determined prior to use in the study by comparing weights of soil before and after oven-drying at ca. 100 °C.
PROPERTIES - Calke sandy loam
- Soil texture: Sand: 69 %; silt: 18 %, clay: 13 %
- Soil taxonomic classification: Sandy Loam
- Soil classification system: USDA
- pH: 1:1 Soil:H2O Ratio: 5.8; 1N KCl: 5.4; 0.01M CaCl2 (1:2): 5.6
- Organic carbon (%): 3.13 (organic matter/1.724)
- CEC (meq/100 g): 12.1
- Moisture content (% w/w): 14.78
- Bulk density (disturbed) (g/cm^3): 1.12
- Water holding capacity: pF 2.0 – WHC 0.1 bar: 20.9; pF 2.5 – WHC 0.33 bar: 15.7
PROPERTIES - South Witham Clay Loam
- Soil texture: Sand: 41%; silt: 26 %, clay: 33 %
- Soil taxonomic classification: Clay Loam
- Soil classification system: USDA
- pH: 1:1 Soil:H2O Ratio: 7.3; 1N KCl: 6.9; 0.01M CaCl2 (1:2): 7.2
- Organic carbon (%): 3.71 (organic matter/1.724)
- CEC (meq/100 g): 25.6
- Moisture content (% w/w): 22.05
- Bulk density (disturbed) (g/cm3): 1.13
- Water holding capacity: pF 2.0 – WHC 0.1 bar: 27.2; pF 2.5 – WHC 0.33 bar: 23.2
PROPERTIES - Lockington Sandy Clay Loam
- Soil texture: Sand: 47 %; silt: 26 %, clay: 33 %
- Soil taxonomic classification: Clay Loam
- Soil classification system: USDA
- pH: 1:1 Soil:H2O Ratio: 5.7; 1N KCl: 5.1; 0.01M CaCl2 (1:2): 5.6
- Organic carbon (%): 3.07 (organic matter/1.724)
- CEC (meq/100 g): 22.9
- Moisture content (% w/w): 22.55
- Bulk density (disturbed) (g/cm3): 1.12
- Water holding capacity: pF 2.0 – WHC 0.1 bar: 28.2; pF 2.5 – WHC 0.33 bar: 24.4
PROPERTIES - Hagan Loamy Sand
- Soil texture: Sand: 87 %; silt: 8 %, clay: 5 %
- Soil taxonomic classification: Loamy Sand
- Soil classification system: USDA
- pH: 1:1 Soil:H2O Ratio: 5.7; 1N KCl: 5.3; 0.01M CaCl2 (1:2): 5.5
- Organic carbon (%): 2.90 (organic matter/1.724)
- CEC (meq/100 g): 10.1
- Moisture content (% w/w): 11.33
- Bulk density (disturbed) (g/cm3): 1.36
- Water holding capacity: pF 2.0 – WHC 0.1 bar: 11.5; pF 2.5 – WHC 0.33 bar: 6.1 - Details on test conditions:
- EXPERIMENTAL APPARATUS
The vessels used throughout the study were PTFE tubes of approximately 50 mL capacity and sealed with a PTFE screw cap, with the exception of the preliminary apparatus check which used both PTFE tubes and thick walled, plastic coated glass tubes of approximately 100 mL capacity and the exception of the soil: solution ratio determination where only glass tubes were used. All tubes were weighed before and after addition of soil (where appropriate) and then after addition of calcium chloride (CaCl2) solution. Each tube was uniquely labelled.
The concentration of calcium chloride (CaCl2) solution used throughout the study was 0.01 M and the solutions were all prepared using the dihydrate and the pH measured before use. All sample tubes were shaken in the dark, on an end-over-end shaker (unless otherwise specified), at a temperature of 20 ± 1 °C. Centrifugation was performed at 3 400 rpm (2 390 rcf) and ca. 20 °C.
PREPARATION AND APPLICATION OF TEST SOLUTIONS
- Preparation of Test Solutions
The test material, nominally 3.1 mg of [14C]-labelled test material was diluted to approximately 10 mL with 2 mL acetonitrile and 8 mL of de-ionised water to give the treatment stock solution. A 200 μL aliquot of this solution was taken and diluted accurately to 10 mL with acetonitrile and 100 μL aliquots were removed and counted by LSC to determine the exact concentration. The concentration of this stock solution was determined to be 0.36 mg mL^-1.
Treatment solutions for each phase of the preliminary and definitive studies were prepared by removing a suitable volume of the stock solution and diluting to the required volume with 0.01 M calcium chloride solution (as described below). The treatment solutions were prepared to ensure that the organic solvent content in any phase of the study would not exceed 0.1 %.
- Application Procedures
Aliquots of the prepared treatment solutions of [14C]-test material were dispensed using a calibrated pipette into sample tubes containing a pre-equilibrated slurry of soil and calcium chloride solution to achieve the target ratio and nominal concentrations as described in the following section. The exception was the preliminary experiment to measure adsorption to tubes, where the appropriate treatment solution was weighed directly into the tubes.
The dpm and µg applied to each tube were determined by performing dose checks during treatment. Aliquots of 1 mL of the appropriate treatment solution were applied directly to scintillation vials and counted by LSC, with the exception of the highest concentration in the definitive tests where the 1 mL treatment was diluted 10-fold prior to LSC analysis.
EXPERIMENTAL CONDITIONS
- Preliminary Tests
Preliminary investigations were carried out to check for adsorption to the tubes, to determine any background radioactivity in the soils, to determine the soil: solution ratio to be used, to check the stability of the test material in 0.01 M calcium chloride and to determine the time required for the compound to equilibrate between soil and water under both adsorption and desorption conditions.
The adsorption and desorption tests were performed using sterilised soils.
- Adsorption to the Tubes
A 14 μL aliquot of the [14C]-test material stock solution was added to approximately 500 mL of calcium chloride solution to give a treatment solution at a concentration of ca. 0.01 mg L^-1. Aliquots of this treatment solution were weighed for analysis by LSC. Approximately 75 mL of the treatment solution was added to two glass tubes and 40 mL was added to the two PTFE tubes, the tubes were tightly capped, weighed and shaken for approximately 24 hours. Further aliquots of the solutions were then removed from each tube for analysis by LSC. This test was performed on both glass and PTFE tubes to check which tubes would be best to use for the remaining phases of the study.
- Soil Background Control
Approximately 4 g oven dried equivalent (ode) of each soil was added to each of two tubes. Approximately 20 mL of calcium chloride solution was added to each tube and these were shaken for 24 hours. The tubes were then removed, weighed and centrifuged for 10 minutes. Accurately weighed aliquots of the supernatant were analysed by LSC.
- Soil: Solution Ratio Determination
A treatment solution was prepared to allow treatment at a nominal concentration of 0.25 mg L^-1 by diluting 0.55 mL of the stock solution to 20 mL with calcium chloride solution. For each soil, 8, 13.3 and 20 g (ode) portions were added to single, uniquely numbered adsorption/desorption tubes. An appropriate volume of calcium chloride solution (39 mL less the moisture content of the soil) was added to each tube to give soil: solution ratios of approximately 1:5, 1:3 and 1:2 and the tubes were shaken overnight to pre-equilibrate (ca. 16 hours) prior to treatment. Following pre-equilibration, 1 mL of the treatment solution was added to each tube. The tubes were capped and then shaken for 24 hours. The tubes were then removed, centrifuged for 10 minutes, the supernatant decanted and the tubes reweighed. Weighed aliquots of the supernatant were taken for analysis by LSC. All supernatants were analysed by HPLC.
HPLC analysis indicated significant breakdown of the test material so all soils under test were sterilised by gamma irradiation. All tests from the adsorption equilibrium onwards were performed using sterilised soils.
- Stability in 0.01 M Calcium Chloride
In order to assess the stability of the test material in the test medium, a single tube containing 0.01 M calcium chloride (ca. 40 mL) without soil was prepared and treated with 1 mL of the treatment solution from the soil: solution ratio determination experiment as outlined above. This solution was then analysed periodically by HPLC.
- Adsorption Equilibrium Time Determination
A treatment solution was prepared to allow treatment at a nominal concentration of 0.5 mg L^-1 by diluting 0.55 mL of the stock solution to 20 mL with calcium chloride solution. Five portions of soil (6.7 g ode for the Calke and the South Witham soil, 4 g ode for the Lockington soil and 10 g ode for the Hagen (Refesol 04-A) loamy sand soil) were weighed into uniquely numbered tubes. Calcium chloride solution (19 mL less the moisture content of the soil) was added and the mixture was placed on an end-over-end shaker to pre-equilibrate (ca. 16 hours) prior to treatment. Following pre-equilibration, 1 mL of the treatment solution was added to each tube and the tubes were placed back on an end-over-end shaker. One tube from each soil type was removed after ca. 2, 4, 24, 48 and 72 hours. At each time point, the tubes were centrifuged for 10 minutes, the supernatants removed and the tubes reweighed. Weighed aliquots of the supernatant were taken for analysis by LSC.
Following the adsorption phase, the soils were extracted using 20 mL portions of acetonitrile followed by acetonitrile: water (4:1 v/v). The tubes were placed on a wrist action shaker for 30 minutes. The tubes were then removed, centrifuged for 10 minutes and the supernatants transferred to a pre-weighed plastic bottle. All extracts were combined, weighed and aliquots of each supernatant were removed in order to determine the radioactivity by LSC. Selected adsorption supernatants and combined solvent extracts were analysed by HPLC.
- Desorption Equilibrium Time Determination
Further tubes (4 per soil) were prepared and treated as for the preliminary adsorption equilibrium time determination. Following pre-equilibration and treatment, the tubes were shaken for 48 hours. The tubes were centrifuged for 10 minutes, the supernatants were removed by decantating and the tubes reweighed. An additional portion of fresh calcium chloride solution was added to each tube (approximately equal to that removed), which was re-capped, reweighed and then placed on an end-over-end shaker. One tube was removed for each soil after approximately 1, 2, 24 and 48 hours. At each time point, the tubes were weighed, centrifuged for 10 minutes, the supernatants removed and the tubes reweighed. Aliquots of each supernatant were weighed and the radioactivity determined by LSC.
In order to prove extractability and parental mass balance for the planned duration of the definitive phase, the 2 hour desorption equilibrium samples were extracted as for the adsorption equilibrium time determination using 30 mL for each extract.
- Definitive Tests
The definitive test was conducted using sterilised soils. All solutions were shaken in the dark at a temperature of 20 ± 1 °C. Following consultation with the sponsor, a 72 hour adsorption equilibrium time was adopted for the definitive phase.
- Adsorption Phase
The experimental conditions for this phase of the study for all soils are summarised below:
- Condition of the soil Soils were sieved to ≤ 2 mm. Before application soils were preequilibrated with 0.01 M CaCl2 solution overnight.
- Soil (weight/replicate): 10 g ode for Calke, South Witham and Hagen (Refesol 04-A) loamy sand soils and 6.7 g ode for the Lockington soil. Each test was performed in duplicate.
- Equilibrium solution used 0.01 M aqueous CaCl2 solution.
- Control used: Controls without soil or test material were used in the preliminary studies only.
- Nominal application rates: 1.0, 0.4, 0.1, 0.04 and 0.01 mg L^-1
- Analytically measured concentrations: 1.03, 0.41, 0.10, 0.040, 0.011 mg L^-1
- Identity and concentration of co-solvent: ≤ 0.1% v/v acetonitrile.
- Soil: solution ratio: 1:3 for Lockington (6.7 g soil to 20 mL solution) and 1:2 for all other soils (10 g soil to 20 mL solution).
- Initial pH of the equilibration solution: a) pH of 0.01M CaCl2 solution without soil 6.38 (Batch 3); b) pH with soil but without test material 6.41 - 7.29 (control samples).
- Number of replications (control and treatment): Duplicate
- Equilibration: Time: Pre-equilibration overnight (ca. 16 hours); Temperature: 20 ± 1 °C; Dark: yes; Shaking method: End-over-end shaker; Shaking time: 72 hours (adsorption).
- Method of separation of supernatant: Centrifugation
- Centrifugation: Speed: 3400 rpm; Duration: 10 minutes; Method of separation of soil and solution: Supernatant was carefully decanted.
Treatment solutions were prepared by dispensing 1.38 mL, 550, 138, 55 and 14 μL of the stock solution and diluting to 25 mL with calcium chloride solution. Uniquely labelled duplicate tubes were prepared for each soil at each of five concentrations. Approximately 10 g ode portions of the Calke, South Witham and Hagen (Refesol 04-A) loamy sand soil and 6.7 g ode portions of the Lockington soil were weighed into pre-weighed tubes. The tubes were capped and re-weighed. Calcium chloride solution (19 mL minus the soil moisture content) was added and the mixture was shaken for ca. 16 hours to pre-equilibrate prior to treatment. Following pre-equilibration, 1 mL of the appropriate treatment solution was added to each tube to allow treatment at nominal concentrations of 1.0, 0.4, 0.1, 0.04 and 0.01 mg L^-1 [14C]-test material.
The soil solutions were mixed and slurried for 72 hours on an end-over-end shaker. The tubes were removed from the shaker, weighed and centrifuged for 10 minutes. The supernatant solutions were removed by decantating and the tubes containing the soil pellets were reweighed. Aliquots of each supernatant were weighed and the radioactivity determined by LSC.
- Desorption Phase
The experimental conditions for this phase of the study are summarised for all soils below:
- Were the soil residues from the adsorption phase used: Yes
- Amount of test material present in the adsorbed state/adsorbed amount: 17.7 – 40.6 % depending on soil type and concentration.
- Number of desorption cycles: One
- Equilibrium solution and quantity used per treatment for desorption: The decanted solution was replaced by an approximately equal volume of fresh 0.01 M aqueous CaCl2 solution.
- Soil: solution ratio: 1:3 for Lockington, 1:2 for all other soils.
- Number of replications: Controls: None; Treatments: Duplicate.
- Equilibration: Time: 2 hours; Temperature: 20 ± 1 °C; Dark: Yes; Shaking method: End-over-end shaker.
- Method of separation of supernatant: Centrifugation
- Centrifugation: Speed: 3400 rpm; Duration: 10 minutes; Method of separation of soil and solution: Supernatant was carefully decanted.
- Solvent extraction: Number of extractions: 1 x 20 mL for all soil samples; Solvent: Acetonitrile; Time: 30 minutes; Temperature: 20 ± 1 °C; Dark: Yes; Shaking method: End-over-end shaker.
Following removal of the adsorption supernatant, an additional portion of fresh calcium chloride solution (approximately equal to the volume removed) was added to each tube, which was capped and weighed. Each tube was placed on an end-over-end shaker. After approximately 2 hours, the tubes were removed, weighed and centrifuged for 10 minutes. The supernatant (desorbate) was removed and the tubes were reweighed. Aliquots of each supernatant were weighed and the radioactivity determined by LSC.
- Extraction
Following desorption, all tubes were solvent extracted. Approximately 20 mL of acetonitrile was added to each tube and the tubes weighed. The tubes were placed on the end-over-end shaker for ca. 30 minutes, removed, reweighed, centrifuged for 10 minutes and the supernatants decanted. Following removal of the supernatant, the tube and soil pellet were reweighed to enable the weight of each supernatant to be calculated.
Aliquots of each supernatant were weighed and the radioactivity determined by LSC. All tubes, containing soil, were then allowed to air dry, reweighed without the cap prior to homogenisation and combustion.
- Stability
All adsorption and desorption supernatants, plus solvent extracts from top concentration of the definitive phase were analysed by HPLC. Sufficient data was obtained to demonstrate the stability of the [14C]-test material for the total duration of the definitive phase.
- Mass Balance
All supernatants, extracts and soil residues were assayed by LSC to determine the total recovery of radioactivity. The mass balance was calculated by comparison to the applied radioactivity as determined from the appropriate dose check. - Duration:
- 72 h
- Initial conc. measured:
- >= 0.01 - <= 1 other: mg/L^-1
- Temp.:
- 20 °C
- Remarks:
- All soil types
- Computational methods:
- SAMPLE CALCULATIONS
The weights and radiocarbon content (dpm) data for each sample were transferred directly into a laboratory information management system (LIMS). Balance weights were automatically transferred using the program WinWedge. DPM values were transferred using the program Yukon from the LSC to the LIMS. The LIMS then used the data to calculate solution and soil concentrations and radiochemical recoveries using validated equations. The soil and solution concentrations were exported to Microsoft Excel spreadsheets to generate the adsorption and desorption isotherms and calculate the Freundlich coefficients.
SOIL CONCENTRATION - ADSORPTION
The amount of [14C]-test material adsorbed to the soil is calculated as the difference between the amount applied (μg) and that found in the aqueous phase. As a quantity of aqueous solution (residual water) remains in the soil pellet following centrifugation, an allowance is made for this. The amount of residual water is calculated using the equation:
R1 = Ws - Wode
Where:
R1 = Residual water 1 (g)
Ws = Weight of soil pellet after removal of supernatant (g)
Wode = Weight of oven dried equivalent soil (g)
The concentration of test material in the residual water is assumed to be equal to that in the supernatant. The concentration of test material adsorbed to the soil is therefore calculated from the equation:
Cs1 = [C1MT - (Cw1M1 + Cw1R1)] / Wode
Where:
Cs1 = Soil concentration after adsorption (μg g^-1)
C1 = Concentration of treatment solution (μg g^-1)
Cw1 = Concentration of adsorption supernatant (μg g^-1)
MT = Weight of treatment (g)
M1 = Weight of adsorption supernatant decanted (g)
R1 = Residual water (adsorption phase) (g)
Wode = Oven dried soil equivalent (g)
SOIL CONCENTRATION - DESORPTION
The concentration on the soil can be calculated from the difference between the total amount of test material in the tube at the start of the cycle and the amount in the aqueous phase. As with the adsorption phase an allowance was made for the test material in the residual water.
Csx = [(Cs(x-1)Wode + Cw(x-1)R(x-1)) - (CwxMx + CwxRx)] / Wode
and Rx = Wx- Wode
Where:
Csx = Soil concentration (μg g^-1)
Cs(x-1) = Soil concentration (μg g-1) previous cycle
Cwx = Concentration in aqueous phase at equilibrium (μg g^-1)
Cw(x-1) = Concentration in aqueous phase, previous cycle (μg g^-1)
Rx = Residual water after this desorption cycle (g)
R(x-1) = Residual water (after previous cycle) (g)
Mx = Weight of supernatant decanted (g)
Wode = Oven dried equivalent weight of soil (g)
Wx = Weight of soil pellet after decanting supernatant (g)
FREUNDLICH ISOTHERMS (Kf, Kdes)
The Freundlich adsorption constant (Kf) and (1/n) for each soil, can be calculated from the Freundlich equation.
Cs1 = KfCw1^(1/n)
Where:
Cs1 = Soil concentration after adsorption (μg g^-1)
Cw1 = Concentration of adsorption supernatant (μg g^-1)
1/n = constant
The constants are obtained by plotting log Cs1 vs. log Cw1 to give 1/n as the slope and log Kf as the intercept.
For the desorption cycle, the desorption coefficient, Kdes, was determined using the Freundlich equation.
Csx = KdesCwx^(1/n)
Where:
Csx = concentration adsorbed to soil at equilibrium (μg g^-1)
Cwx = concentration in solution at equilibrium (μg g^-1)
1/n = constant
SORPTION COEFFICIENTS NORMALISED FOR PERCENT ORGANIC CARBON (Koc, Kocdes)
The Freundlich constant was expressed as a function of the organic carbon content of the soil using the equation:
Koc = (Kf x 100) / % organic carbon
In addition, the desorption coefficient was expressed as a function of organic carbon content of the soil (Kocdes), using the equation:
Kocdes = (Kdes x 100) / % organic carbon
STATISTICAL METHODS
Where a mean or average value is quoted it is the arithmetical mean which is used:
Average = xmean = (x1 + x2 + x3 + ….xn) / n
The standard deviation (SD) from the mean, σ(n-1) is calculated from:
SD = σ(n-1) = √[(x1 - xmean)^2 + (x2 - xmean)^2 + … (xn - mean)^2] / n-1
This calculation assumes that only a sample of the population of x values is known and was performed in Excel (version 2007) using the STDEV function.
The coefficient of variation, cv, is calculated from:
Cv = (σ(n-1) / xmean) x 100 % - Key result
- Type:
- Koc
- Value:
- >= 12 - <= 34 other: mL g^-1
- Temp.:
- 20 °C
- Matrix:
- Soil
- % Org. carbon:
- >= 2.9 - <= 3.71
- Remarks on result:
- other: pH range: 5.5 - 7.2
- Key result
- Type:
- Kd
- Value:
- >= 24 - <= 54 other: mL g^-1
- Temp.:
- 20 °C
- Matrix:
- Soil
- % Org. carbon:
- >= 2.9 - <= 3.71
- Remarks on result:
- other: pH range: 5.5 - 7.2
- Adsorption and desorption constants:
- - Adsorption Measurements
In the definitive adsorption test, the amount of applied test material adsorbed ranged from 19.3 to 33.7 % in the Calke, 17.7 to 26.8 % in the South Witham, 17.8 to 30.6 % in the Lockington and 32.1 to 40.6 % in the Hagen.
For all soils the fit of log Cs1 vs. log Cw1 to a linear equation was good with correlation coefficients of 0.99 to 1.00.
The Kf values ranged from 0.46 mL g^-1 in the South Witham soil to 0.98 mL g^-1 in the Hagen soil.
The relationship between the soil and solution concentration was non- linear for all of the soils tested, with 1/n values ranging from 0.852 to 0.926.
The values for Koc ranged from 12 mL g^-1 in the South Witham soil to 34 mL g^-1 in the Hagen soil, with a mean value of 21 mL g^-1, indicating that the test material can be classified as being mobile, according to the Briggs classification and of very high mobility according to the McCall classification.
- Desorption Measurements
At the end of the desorption phase, the amount of test material desorbed, expressed as a percentage of the initial amount adsorbed, ranged from 31.0 to 34.9 % in the Calke, 32.1 to 35.4 % in the South Witham, 33.4 to 38.4 % in the Lockington and 33.9 to 38.3 % in the Hagen.
For all soils the fit of log Cs1 vs. log Cw1 to a linear equation was good with correlation coefficients of 0.99 to 1.00.
Kdes values obtained ranged from 0.88 mL g^-1 in the South Witham soil to 1.55 mL g^-1 in the Hagen soil. The values of 1/n for the desorption cycle were very similar to those obtained for the adsorption for each soil tested.
The Kocdes values ranged from 24 mL g^-1 in the South Witham soil to 54 mL g^-1 in the Hagen soil, with a mean value of 37 mL g^-1. - Concentration of test substance at end of adsorption equilibration period:
- The concentrations in the soil and water phases and the percent of applied adsorbed after the adsorption phase are given below.
- Concentration of test substance at end of desorption equilibration period:
- The concentrations in the soil and water phases and the percent of applied adsorbed after the desorption phase are given below.
- Sample no.:
- #1
- Duration:
- 72 h
- % Adsorption:
- >= 19.3 - <= 33.7
- Remarks on result:
- other: Calke sandy loam
- Sample no.:
- #2
- Duration:
- 72 h
- % Adsorption:
- >= 17.7 - <= 26.8
- Remarks on result:
- other: South Witham clay loam
- Sample no.:
- #3
- Duration:
- 72 h
- % Adsorption:
- >= 17.8 - <= 30.6
- Remarks on result:
- other: Lockington sandy clay loam
- Sample no.:
- #4
- Duration:
- 72 h
- % Adsorption:
- >= 32.1 - <= 40.6
- Remarks on result:
- other: Hagen loamy sand
- Sample no.:
- #1
- Duration:
- 2 h
- % Desorption:
- >= 31 - <= 34.9
- Remarks on result:
- other: Calke sandy loam
- Sample no.:
- #2
- Duration:
- 2 h
- % Desorption:
- >= 32.1 - <= 35.4
- Remarks on result:
- other: South Witham clay loam
- Sample no.:
- #3
- Duration:
- 2 h
- % Desorption:
- >= 33.4 - <= 38.4
- Remarks on result:
- other: Lockington sandy clay loam
- Sample no.:
- #4
- Duration:
- 2 h
- % Desorption:
- >= 33.9 - <= 38.3
- Remarks on result:
- other: Hagen loamy sand
- Transformation products:
- not measured
- Details on results (Batch equilibrium method):
- PHYSICAL CONDITIONS
The test systems were maintained in the dark and at a temperature of 20 ± 1 °C throughout the study. The pH of the calcium chloride solutions used during this study was found to be in the range 6.32 to 6.42.
ANALYTICAL METHODOLOGY
- Column Recoveries
HPLC column recoveries were good with values of 99.4 and 95.4 %.
- Confirmation of Compound Stability
The test material was found to be stable in 0.01 M calcium chloride for at least 6 days, indicating stability for greater than the duration of the definitive phase of the study. Selected supernatants and solvent extracts were examined by HPLC during the preliminary adsorption equilibrium time determination.
The adsorption equilibrium determination proved greater than 90 % of [14C]- test material was extracted from the soil after 48 hours of adsorption. [14C]-test material accounted for greater than 97 % of the region of interest in the HPLC radiochromatogram for all samples run during the preliminary and definitive study. It was concluded that no significant degradation of [14C]-test material occurred over the duration of the study. Therefore no adjustment to the adsorption or desorption coefficients was necessary.
PRELIMINARY TESTS
- Adsorption to Tubes
Recovery of radioactivity from the tubes was quantitative with a mean recovery of 99.6 %. No correction for adsorption to tubes was therefore required; PTFE tubes chosen to be used for the remaining phases of the study as they are practically easier to use than glass tubes.
- Soil Background Control
The level of background radioactivity detected was negligible in all soils and therefore no correction was necessary.
- Soil: Solution Ratio
[14C]-test material indicated adequate adsorption (within the preferred 20 - 80 % limit) to the soil for all selected soil: solution ratios tested, with the exception of the Hagen soil (Refesol 04-A) at the 1:5 ratio. A soil: solution ratio of 1:2 was selected for the Calke, South Witham and Hagen (Refesol 04-A) loamy sand soils (55.4, 58.7 and 36.4 % adsorbed, respectively) and a 1:3 soil: solution ratio was selected for the Lockington soil (64.4 % adsorbed).
- Adsorption Equilibrium Time
The adsorption equilibrium of test material was reached after 48 hours for all soils, with the exception of the Hagen (Refesol 04-A) loamy sand soil where the adsorption continued to increase between 48 hours and 72 hours. Parental mass balances were determined by LSC and HPLC analysis of the adsorption supernatants and solvent extracts. Following a 48-hour adsorption period, 93.6 % of applied radioactivity was confirmed as test material in the Calke sandy loam, 90.0 % in the South Witham clay loam, 92.1 % in the Lockington sandy clay loam and 92.6 % in the Hagen (Refesol 04-A) loamy sand.
An adsorption equilibrium time of 48 hours was selected for the desorption equilibrium phase of the study. Following consultation with the sponsor, a 72-hour adsorption time was later selected for the definitive phase of the study. A pre-equilibration time of ca. 16 hours was used.
- Desorption Equilibrium Time and Parental mass Balance
In the desorption equilibrium test there was very little change in the levels of radioactivity in solution between 2 hours and 48 hours in any of the soils tested. HPLC analysis of the 2 hour desorption supernatants, solvent extracts and corresponding adsorption supernatants indicated no significant degradation, with > 97 % of the chromatogram attributable to the test material in each case. A desorption time of 2 hours for all soils was selected for the definitive phase.
- Definitive Phase
An adsorption equilibrium time of 72 hours and a desorption equilibrium time of 2 hours were selected for all soils. A pre-equilibration time of ca. 16 hours was used. An additional test was performed alongside the definitive test using the South Witham soil without pre-equilibration with calcium chloride. As this presented no significantly different results to the test with pre-equilibration, the results were excluded from any calculations of mean values.
- Material Balance
The recovery of radioactivity was quantitative, with all recoveries within the acceptable range of 90 - 110 % of applied radioactivity. The overall material balance for individual samples was in the range of 94.3 - 99.2 % for the Calke sandy loam (mean 96.8 %), 97.1 - 101.0 % for the South Witham clay loam (mean 98.3 %), 97.6 - 105.8 % for the Lockington sandy clay loam (mean 99.9 %), and 94.9-99.3 % for the Hagen (Refesol 04-A) loamy sand (mean 97.0 %).
CONFIRMATION OF COMPOUND STABILITY/IDENTITY
- Test Material Identity and Purity
The radiopurity of the treatment solution was determined as 97.87 % by HPLC.
Comparison of LC/MS data with that of a certified reference standard confirmed the identity of the test material.
- Test Material Stability during the Definitive Test
The stability of the test material for the total duration of the definitive test, consisting of a 72 hour adsorption cycle and a 2 hour desorption cycle, was demonstrated during the definitive study.
- Storage Stability
All samples were analysed within 3 days of generation.
CONCLUSIONS
- Major Outcomes of Study
The Koc values obtained ranged from 12 to 34 mL g^-1 (mean 21 mL g^-1). Freundlich exponents were non-linear for all of the soils, with 1/n values of 0.85 to 0.93, indicating significant change in the relationship between the amount adsorbed onto the soil and the amount in solution through the concentration range.
The Kocdes values ranged from 24 to 54 mL g^-1 (mean 37 mL g^-1).
- Significance of Results to Environmental Behaviour of Test Material
The determined Koc values indicate that the test material can be classified as being mobile in soil according to the Briggs classification and as having a very high mobility in soil according to the McCall classification. - Validity criteria fulfilled:
- yes
- Conclusions:
- Under the conditions of this study, the mean determined Koc and Koc des values were 21 and 37 mL g^-1 respectively, with corresponding mean 1/n values of 0.881 and 0.900.
- Executive summary:
The adsorption and desorption characteristics of the test material were studied in four soils using the batch equilibrium method in accordance with the standardised guidelines OECD 106 and US EPA OPPTS 835.1230 under GLP conditions.
The soils used were Calke sandy loam (UK, pH 5.6, 3.13 % organic carbon), South Witham clay loam (UK, pH 7.2, 3.71 % organic carbon), Lockington sandy clay loam (UK, pH 5.6, 3.07 % organic carbon) and Hagen (Refesol 04-A) loamy sand (Germany, pH 5.5, 2.90 % organic carbon). Due to significant breakdown of the test material observed in the soil: solution ratio preliminary test, subsequent tests were performed using soils that had been sterilised by gamma irradiation.
An additional test was carried out during the definitive phase using the South Witham soil, which was treated without pre-equilibration with calcium chloride. As this test showed no significant differences in adsorption, it was excluded from all mean data.
The adsorption phase of the study was carried out using sterile, pre-equilibrated soils with [14C]-test material at concentrations of approximately 1.0, 0.4, 0.1, 0.04 and 0.01 mg L^-1 in the dark and at 20 ± 1 °C for 72 hours for all soils. The equilibration solution used was 0.01 M aqueous CaCl2, with a soil: solution ratio of 1:2 for the Calke sandy loam, the South Witham clay loam and for the Hagen (Refesol 04-A) loamy sand and a 1:3 ratio for the Lockington sandy clay loam. The desorption phase of the study was carried out for 2 hours with fresh 0.01 M aqueous CaCl2 applied to pre-adsorbed soil for one desorption cycle.
The aqueous supernatant after adsorption and desorption was separated by centrifugation and analysed by LSC. After desorption, the soil was extracted with acetonitrile and following extraction, the soil was combusted and the trapped CO2 analysed by LSC. The adsorption parameters were calculated using the Freundlich isotherm.
It was proven during the preliminary tests that greater than 90 % of applied radioactivity could be extracted from the soil after 48 hours adsorption and close to 90 % extractable after 72 hours. In both the preliminary and definitive tests, [14C]-test material accounted for greater than 97 % of the region of interest in all supernatants and extracts analysed by HPLC. It was concluded therefore that no significant degradation of [14C]-test material occurred over the duration of the study.
The overall material balances in the definitive phase were determined by LSC of the supernatants after adsorption, desorption and solvent extraction and combustion of the remaining soils. The overall material balances were 96.8 % for the Calke sandy loam, 98.3 % for the South Witham clay loam, 99.9 % for the Lockington sandy clay loam and 97.0 % for the Hagen (Refesol 04-A) loamy sand.
In the definitive adsorption test, the amount of applied test material adsorbed ranged from 19.3 to 33.7 % in the Calke sandy loam, 17.7 to 26.8 % in the South Witham clay loam, 17.8 to 30.6 % in the Lockington sandy clay loam and 32.1 to 40.6 % in the Hagen (Refesol 04-A) loamy sand.
The calculated adsorption constants (Kf) of the Freundlich isotherms for the soils ranged from 0.46 mL g^-1 in the South Witham clay loam to 0.98 mL g^-1 in the Hagen (Refesol 04-A) loamy sand. The Freundlich exponents (1/n) ranged from 0.852 in the Calke sandy loam to 0.926 in the Hagen (Refesol 04-A) loamy sand, indicating that the concentration of the test material does have a significant effect on the adsorption behaviour in the examined concentration range.
The adsorption Koc values ranged from 12 mL g^-1 in the South Witham clay loam to 34 mL g^-1 in the Hagen (Refesol 04-A) loamy sand.
At the end of the desorption phase, the amount of test material desorbed, expressed as a percentage of the initial amount adsorbed, ranged from 31.0 to 34.9 % in the Calke sandy loam, 32.1 to 35.4 % in the South Witham clay loam, 33.4 to 38.4 % in the Lockington sandy clay loam and 33.9 to 38.3 % in the Hagen (Refesol 04-A) loamy sand.
The desorption Kdes values ranged from 0.88 mL g^-1 in the South Witham clay loam to 1.55 mL g^-1 in the Hagen (Refesol 04-A) loamy sand.
For all soils the data fit to a linear equation was good for both adsorption and desorption with correlation coefficients of 0.99 to 1.00.
Under the conditions of this study, the mean determined Koc and Koc des values were 21 and 37 mL g^-1 respectively, with corresponding mean 1/n values of 0.881 and 0.900. Thus, according to Briggs, the test material can be classified as being mobile in soil and, according to the McCall classification, as having very high mobility in soil.
- Endpoint:
- adsorption / desorption, other
- Type of information:
- experimental study
- Adequacy of study:
- supporting study
- Reliability:
- 2 (reliable with restrictions)
- Rationale for reliability incl. deficiencies:
- comparable to guideline study with acceptable restrictions
- Reason / purpose for cross-reference:
- reference to same study
- Qualifier:
- according to guideline
- Guideline:
- other: OECD Test guideline 'Adsorption-Desorption'.
- Deviations:
- not specified
- GLP compliance:
- no
- Type of method:
- other: Not specified
- Media:
- soil
- Radiolabelling:
- yes
- Test temperature:
- 20 °C
- Analytical monitoring:
- no
- Matrix no.:
- #1
- Matrix type:
- sandy loam
- % Clay:
- 15.8
- % Silt:
- 18.4
- % Sand:
- 63.4
- pH:
- 6.9
- Matrix no.:
- #2
- Matrix type:
- sandy loam
- % Clay:
- 13.2
- % Silt:
- 28.6
- % Sand:
- 55.5
- pH:
- 6.7
- Matrix no.:
- #3
- Matrix type:
- other: Coarse sandy soil: 0 - 25 cm depth
- % Clay:
- 4
- % Silt:
- 6.6
- % Sand:
- 86.9
- pH:
- 6.6
- Details on matrix:
- COLLECTION AND STORAGE
- Geographic location: Soils were from three Danish Experimental Farms (#1 Flakkebjerg, #2 Roskilde and #3 Jyndevad, providing sandy loam, sandy loam and coarse sandy soil, respectively). The soils were grown with rotational crops including cereals.
- Sampling depth: Subsoil was from undisturbed cores of coarse sandy soil sampled 33 - 66 and 66 - 99 cm below soil surface (Jyndevad). Adsorption coefficients were calculated for the soil at a depth of 0 - 25 cm. The depth for both Flakkebjerg and Roskilde was 0 - 30 cm
- Storage conditions: Stored at 5 °C with about 25 % of total water holding capacity (WHC) until use. Subsoil was kept in stainless steel tubes at 5 °C.
- Storage length: Storage for less than one month.
- Soil preparation: After sampling, the surface soils were sieved (< 2 mm).
PROPERTIES OF THE SOILS (in addition to defined fields)
- WHC (Water holding capacity, % of wet soil):
Flakkebjerg: 31
Roskilde: 31
Jyndevad: 29
- Humus (%C x 1.72):
Flakkebjerg: 2.4
Roskilde: 2.6
Jyndevad: 2.4 - Details on test conditions:
- Adsorption (Kd) of the test material was determined by shaking 5 g of soil with 25 mL 0.01 M CaCl2 added to 14C-test material (5 mg L^-1) for 16 h. To avoid degradation the soils were sterilised by irradiation. The soil was precipitated by centrifugation and 14C was counted in aliquots of the supernatant to calculate concentrations of the test material in soil and water. Koc was calculated from Kd and the organic C contents of the soils.
- Sample No.:
- #1
- Duration:
- 16 h
- Initial conc. measured:
- 5 other: mg L^-1
- pH:
- 6.9
- Temp.:
- 20 °C
- Sample No.:
- #2
- Duration:
- 16 h
- Initial conc. measured:
- 5 other: mg L^-1
- pH:
- 6.7
- Temp.:
- 20 °C
- Sample No.:
- #3
- Duration:
- 16 h
- Initial conc. measured:
- 5 other: mg L^-1
- pH:
- 6.6
- Temp.:
- 20 °C
- Sample No.:
- #1
- Type:
- Kd
- Value:
- 0.12 dimensionless
- pH:
- 6.9
- Temp.:
- 20 °C
- Matrix:
- Flakkebjerg soil 0 - 30 cm depth
- Sample No.:
- #2
- Type:
- Kd
- Value:
- 0.2 dimensionless
- pH:
- 6.7
- Temp.:
- 20 °C
- Matrix:
- Roskilde soil 0 - 30 cm depth
- Sample No.:
- #3
- Type:
- Kd
- Value:
- 0.07 dimensionless
- pH:
- 6.6
- Temp.:
- 20 °C
- Matrix:
- Jyndevad soil 0 - 25 cm depth
- Sample No.:
- #1
- Type:
- Koc
- Value:
- 8.4 dimensionless
- pH:
- 6.9
- Temp.:
- 20 °C
- Matrix:
- Flakkebjerg soil 0 - 30 cm depth
- Sample No.:
- #2
- Type:
- Koc
- Value:
- 13.3 dimensionless
- pH:
- 6.7
- Temp.:
- 20 °C
- Matrix:
- Roskilde soil 0 - 30 cm depth
- Sample No.:
- #3
- Type:
- Koc
- Value:
- 5.3 dimensionless
- pH:
- 6.6
- Temp.:
- 20 °C
- Matrix:
- Jyndevad soil 0 - 25 cm depth
- Adsorption and desorption constants:
- The adsorption is low with Kd values between 0.07 and 0.2 and Koc values of 8, 13 and 5, respectively in the three soils. With such low adsorption values the test material is likely to leach if not decomposed in the surface soil.
- Transformation products:
- not measured
- Validity criteria fulfilled:
- not specified
- Conclusions:
- Under the conditions of the study the adsorption of the test material was low with Kd values of 0.07 - 0.2 and Koc values of 8.4, 13.3 and 5.3 in the three soil types.
- Executive summary:
Adsorption of 14C-ring-labelled test material was determined according to the OECD Test Guideline Adsorption - Desorption in three different soil types.
Soils were from three Danish Experimental Farms (#1 Flakkebjerg, #2 Roskilde and #3 Jyndevad, providing sandy loam, sandy loam and coarse sandy soil, respectively).
Adsorption (Kd) of the test material was determined at 20 °C by shaking 5 g of soil with 25 mL 0.01 M CaCl2 added to 14C-test material (5 mg L^-1) for 16 h. To avoid degradation the soils were sterilised by irradiation. The soil was precipitated by centrifugation and 14C was counted in aliquots of the supernatant to calculate concentrations of the test material in soil and water. Koc was calculated from Kd and the organic C contents of the soils.
The adsorption of the test material was low with Kd values of 0.07 - 0.2 and Koc values of 8.4, 13.3 and 5.3 in the three soil types (Flakkebjerg, Roskilde and Jyndevad, respectively). With such low adsorption values the test material is likely to leach if not decomposed in the surface soil.
- Endpoint:
- adsorption / desorption, other
- Type of information:
- experimental study
- Adequacy of study:
- supporting study
- Study period:
- 20 January 1993 to 04 February 1993
- Reliability:
- 2 (reliable with restrictions)
- Rationale for reliability incl. deficiencies:
- guideline study with acceptable restrictions
- Qualifier:
- according to guideline
- Guideline:
- OECD Guideline 106 (Adsorption - Desorption Using a Batch Equilibrium Method)
- Deviations:
- no
- Qualifier:
- according to guideline
- Guideline:
- other: Aanvraag tot toelating van een bestrijdingsmiddel, Toelichting bij het aanvraagformulier A (Dutch Guideline).Bureau Bestrijdingsmiddelen, Postbus 9102, 6700 HC Wageningen, The Netherlands
- Version / remarks:
- June 1991
- Deviations:
- no
- GLP compliance:
- yes
- Type of method:
- batch equilibrium method
- Media:
- soil
- Radiolabelling:
- yes
- Test temperature:
- 20 ± 2 °C
- Analytical monitoring:
- yes
- Matrix no.:
- #1
- Matrix type:
- other: Sandy soil (A)
- % Clay:
- 3.8
- % Silt:
- 7
- % Sand:
- 89.2
- % Org. carbon:
- 5.6
- pH:
- 4.3
- Matrix no.:
- #2
- Matrix type:
- other: Sandy soil (B)
- % Clay:
- 3.8
- % Silt:
- 4.8
- % Sand:
- 91.4
- % Org. carbon:
- 3.6
- pH:
- 4.4
- Matrix no.:
- #3
- Matrix type:
- other: Sandy soil (C)
- % Clay:
- 3.9
- % Silt:
- 6.7
- % Sand:
- 85.4
- % Org. carbon:
- 4.2
- pH:
- 4.3
- Details on matrix:
- COLLECTION AND STORAGE
Three types of soil were used for this investigation.
- Geographic location:
A sandy soil (A) was taken from an undisturbed garden podzol soil in Zeist, The Netherlands (52°06'N, 05°16'E) on 26 December 1992.
Another sandy soil (B) was taken from a permanent grassland near 'De Krakeling' Zeist, The Netherlands (52°06'N, 05°16'E) on 2 January 1993.
A third sandy soil (C) was taken from an arable field near Maarn, The Netherlands (52°04'N, 05°21'E) on 10 January 1993.
- Storage conditions: The soils were stored at 3 ± 2 °C pending use.
- Soil preparation: The soils were sieved (mesh 2 mm) to remove coarse particles before being added to the test flasks. The dry weights were determined by drying subsamples at 105 °C overnight in triplicate.
PROPERTIES
- Organic carbon (%):
Soil A: 5.6 % organic matter (% dry weight)
Soil B: 3.6 % organic matter (% dry weight)
Soil C: 4.2 % organic matter (% dry weight)
- Carbonate as CaCO3:
Soil A: 0.1 % (% dry weight)
Soil B: 0.1 % (% dry weight)
Soil C: 0.1 % (% dry weight) - Details on test conditions:
- The study was carried out to investigate the adsorption of the test material to soil particles of three sandy soil types with a pH (KCl) between 4 and 5. The determinations were carried out by the 'slurry method'. To determine the adsorption constants related to soil organic matter contents (Kom) from the adsorption isotherms, four concentrations of the test material were shaken with soil. The balance of the radioactivity was also determined.
TEST SYSTEM
- Type, size and further details on reaction vessel: The test was carried out in scintillation vials of 20 mL closed with screwcaps.
- Rate of adsorption: One g dry weight of soil was placed into each vial together with 10 mL 0.05 M CaCl2 solution containing 5 mg.L^-1 [14C]-test material. Duplicate samples were taken after 1, 4, 20 and 48 hours (after shaking at 20 ± 2 °C) to determine the equilibrium time of the adsorption. The vials were centrifuged at gmax = 450 for five minutes. Duplicate samples of one mL were pipetted into scintillation vials containing 10 mL of Ultima Gold. A 48 h period was chosen for the adsorption test.
- Adsorption test: One g dry weight of soil was placed in each vial. Appropriate amounts of the stock solution were pipetted into the test vials to reach final concentrations of 5, 10, 20 and 50 μg of test material per vial. CaCl2 solution was added to the vials to make up the volume to 10 mL. The vials were shaken for 48 h at 20 ± 2 °C in a temperature-controlled room. The vials were centrifuged at gmax = 450 for 5 minutes. Duplicate samples of one mL were pipetted into scintillation vials containing 10 mL of Ultima Gold.
- Test performed in closed vessels due to significant volatility of test substance: Scintillation vials of 20 mL were closed with screwcaps.
- Method of preparation of test solution: The vial with [14C]-test material was rinsed 15 times with 1 mL 0.05 M CaCI2 solution. CaCl2 solution was added to a volume of 250 mL. The amount of radioactivity was checked by pipetting a 1 mL aliquot of this solution directly into a scintillation vial containing 10 mL Ultima Gold. The solution contained 5.44 MBq (1.9 mg) test material. 5.9 mg test material was dissolved in 100 mL CaCl2 solution. 10 mL of this solution was pipetted to the solution with [14C]-test material and the total volume was brought to 500 mL. The amount of radioactivity was checked in triplicate. The stock solution contained 5 mg.L^-1 test material.
- Are the residues from the adsorption phase used for desorption: No
RADIOACTIVITY BALANCE
The liquid was pipetted from the soil samples (5 mg.L^-1). The soils were frozen pending analysis. Methanol (10 mL) was added to the soils. The vials were shaken vigorously by hand and centrifuged at gmax = 450 for five minutes. Duplicate samples of one mL were pipetted into scintillation vials containing 10 mL of Ultima Gold. - Sample No.:
- #1
- Duration:
- 48 h
- Initial conc. measured:
- >= 5 - <= 50 other: μg per vial
- Temp.:
- 20 °C
- Remarks:
- All soils
- Sample No.:
- #1
- Type:
- Koc
- Value:
- 4.5 L/kg
- pH:
- 4.3
- Temp.:
- 20 °C
- Matrix:
- Soil A
- Sample No.:
- #2
- Type:
- Koc
- Value:
- 3.5 L/kg
- pH:
- 4.4
- Temp.:
- 20 °C
- Matrix:
- Soil B
- Sample No.:
- #2
- Type:
- Koc
- Value:
- 3.3 L/kg
- pH:
- 4.3
- Temp.:
- 20 °C
- Matrix:
- Soil C
- Adsorption and desorption constants:
- From the data showing the concentration of the test material in the supernatant and adsorbed to the soil after 48 hours, adsorption isotherms were plotted and the adsorption constants calculated according to the Freundlich equation.
Adsorption constants were calculated based on organic matter.
Soil A: K = 4.5 mL/g
Soil B: K = 3.5 mL/g
Soil C: K = 3.3mL/g
Since adsorption was less than 25 % in the 5 mg.L^-1 experiment, a study of the desorption is not necessary according to the guideline. - Recovery of test material:
- More than 94 % of the radioactivity was recovered after the test.
- Transformation products:
- not measured
- Validity criteria fulfilled:
- not specified
- Conclusions:
- Under the conditions of the study adsorption constant values for the test material based on total soil were 4.5, 3.5 and 3.3 mL.g^-1 for sandy soils A, B and C, respectively. Adsorption constants calculated on organic matter base were 80, 97 and 78 mL.g^-1 for sandy soils A, B and C, respectively.
- Executive summary:
The adsorption of the test material to three types of soil was determined essentially according the OECD Guideline 106 and according to the requirements of the Dutch Guidelines and in compliance with GLP.
The study was carried out to investigate the adsorption of the test material to soil particles of three sandy soil types with a pH (KCl) between 4 and 5. The determinations were carried out by the 'slurry method'. To determine the adsorption constants related to soil organic matter contents (Kom) from the adsorption isotherms, four concentrations of the test material were shaken with soil. The balance of the radioactivity was also determined.
Under the conditions of the study adsorption constant values for the test material based on total soil were 4.5, 3.5 and 3.3 mL.g^-1 for sandy soils A, B and C, respectively. Adsorption constants calculated on organic matter base were 80, 97 and 78 mL.g^-1 for sandy soils A, B and C, respectively.
Desorption studies were not carried out because the adsorption of the test material at the 5 mg.L^-1 level was less than 25 %.
- Endpoint:
- adsorption / desorption, other
- Remarks:
- Soil leaching study
- Type of information:
- experimental study
- Adequacy of study:
- supporting study
- Study period:
- 07 July 1995 to 10 May 1996
- Reliability:
- 1 (reliable without restriction)
- Rationale for reliability incl. deficiencies:
- guideline study
- Reason / purpose for cross-reference:
- reference to same study
- Qualifier:
- according to guideline
- Guideline:
- other: FIFRA Pesticide Assessment Guidelines, Subdivision N: Guideline 163-1 (Mobility Studies: Leaching and Adsorption/Desorption Studies)
- Version / remarks:
- 18 October 1982
- Deviations:
- no
- GLP compliance:
- yes
- Type of method:
- other: Column leaching experiments.
- Media:
- soil
- Specific details on test material used for the study:
- Test concentrations were adjusted for purity of the test material and are reported as milligrams of test material per liter of solution (mg/L).
The non-radiolabelled test material was used to isotopically dilute the radiolabelled test material as well as to serve as an analytical reference standard.
TREATMENT OF TEST MATERIAL PRIOR TO TESTING
- Preparation of the test material primary stock solution, purification of the radiolabelled test material and radiopurity analysis: The primary stock solution was prepared by transferring the entire contents of the received vial of [14C]test material via repetitive rinsing with acetonitrile to a final volume of 60 mL. The stock solution was analysed by liquid scintillation counting (LSC) and determined to have a concentration of 0.142 mg/mL (70 660 dpm/μL) based on a specific activity of 224.86 μCi/mg (499 189 dpm/μg; 48.3 mCi/mmol; 8.32 MBq/mg). HPLC-RAM analysis revealed, however, that the [14C]test material had a radiopurity of 94.5 %. Since the determined radiopurity was below 95 %, the [14C]test material primary stock solution was reduced to approximately 1 mL under nitrogen and purified by semi-preparative HPLC. The entire amount of impure [14C]test material was injected (in approximate 200 μL portions) onto a semi-preparative C18 HPLC column and the eluting region, corresponding to test material, was collected and pooled. Most of the acetonitrile (from the mobile phase) was removed under nitrogen, and 30 mL of reagent water was added to the remaining acidic water. After adjusting the pH to approximately 2 with acetic acid, the purified [14C]test material was partitioned with 3 x 50 mL of methylene chloride. The methylene chloride was then removed with nitrogen and the sample was redissolved in 50 mL of acetonitrile. Three 5-μL aliquots of the purified stock solution were profiled by HPLC-RAM, establishing a radiopurity of 100 %. The purified [14C]test material primary stock solution was used to prepare stock solutions for testing. This stock solution was analysed by LSC to have a mean measured concentration of 0.131 mg/mL (total of 6.53 mg [14C]test material), based on the radioactivity and the specific activity.
- Isotopic dilution, determination of specific activity and radiopurity of the [14C]test material primary stock solution: The [14C]test material primary stock solution (43 mL containing 5.633 mg of [14C]test material) was combined with 16.881 mg of non-radiolabelled test material. The mean measured concentration of the isotopically diluted primary stock solution was determined to be 0.527 mg/mL, based on an HPLC-UV assay. The specific activity was determined to be 116 952 dpm/μg (11.3 mCi/mmol), based on a combination of LSC (to quantitate radioactivity), HPLC-UV (to determine the concentration of test material using a calibration curve) and HPLC-RAM (to determine radiopurity). Radiopurity was determined to be 100 %. - Radiolabelling:
- yes
- Test temperature:
- 25 ± 1 °C
- Matrix no.:
- #1
- Matrix type:
- sandy loam
- % Clay:
- 7
- % Silt:
- 27
- % Sand:
- 66
- % Org. carbon:
- 0.9
- pH:
- 7.4
- CEC:
- 14.4 meq/100 g soil d.w.
- Bulk density (g/cm³):
- 1.28
- Matrix no.:
- #2
- Matrix type:
- sandy loam
- % Clay:
- 8
- % Silt:
- 36
- % Sand:
- 56
- % Org. carbon:
- 3.5
- pH:
- 6.3
- CEC:
- 11.9 meq/100 g soil d.w.
- Bulk density (g/cm³):
- 0.94
- Matrix no.:
- #3
- Matrix type:
- sand
- % Clay:
- 6
- % Silt:
- 2
- % Sand:
- 92
- % Org. carbon:
- 1.3
- pH:
- 6.7
- CEC:
- 4.3 meq/100 g soil d.w.
- Bulk density (g/cm³):
- 1.35
- Matrix no.:
- #4
- Matrix type:
- clay
- % Clay:
- 50
- % Silt:
- 34
- % Sand:
- 16
- % Org. carbon:
- 1.6
- pH:
- 7
- CEC:
- 34.9 meq/100 g soil d.w.
- Bulk density (g/cm³):
- 1.09
- Details on matrix:
- SOIL COLLECTION AND STORAGE
Soils were obtained from various agricultural sites within the United States and were intended to represent both light and heavy-textured soils. The sand soil was collected from a site in Macon, Georgia and was received at the Test Facility on 8 June 1995. A sandy loam soil (designated Sandy Loam A) was collected from a site in Timmerman, Washington from the top 2 to 9 inches soil profile and was received at the Test Facility on 9 May 1995. Reportedly, the Washington soil had not received pesticide applications for 3 years prior to collection. Another sandy loam soil (designated as Sandy Loam B) was collected (on 5 June 1995) from a field in Hood River County, Oregon from a depth of 0 to 9 inches and was received at the Test Facility on 7 June 1995. Although an initial non-GLP soil characterisation of the Oregon soil indicated a soil texture corresponding to a silt loam, a subsequent characterisation conducted under GLP regulations indicated that this soil corresponded to a sandy loam. The latter characterisation was used. Reportedly, the Oregon soil had not been treated with agricultural chemicals or fertilizers within 5 years of its collection. A (Sharkey) clay soil was collected on 6 June 1995 from a farm near Watson, Arkansas and was received at the Test Facility on 7 June 1995. The clay soil had been left fallow for the previous ten years.
Prior to use, soils were stored at approximately 4 °C. Before use, the soils were sieved through a 2-mm stainless steel mesh to provide a uniform particle size as specified in the protocol. - Details on test conditions:
- Column Leaching Test
- Preparation of Aged Residues for Column Leaching Test: Two 50-gram portions (dry weight) of each soil type in separate 250-mL Erlenmeyer flasks were fortified with 110 μL of the 0.527 mg/mL [14C]test material primary stock solution resulting in a soil concentration of 1.16 μg/g (ppm). Soils (adjusted to 75 % of FMC prior to dosing except for clay which was dosed dry and then moisturised) were mixed with a stainless steel spatula to evenly distribute test material. After allowing approximately 1 to 2 hours for solvent to evaporate, the soil flasks were connected to trapping trains for the purpose of capturing volatiles. Volatiles were purged from the headspace of soil flasks daily for a period of approximately 30 minutes. Soil samples were incubated in darkness in an environmental chamber at 25 ± 1 °C to their approximate half-life, based on the pilot soil metabolism study data. At the time of harvest, one of the two replicate soil samples per soil type was divided into 3 x 10-gram aliquots (dry weight). One of the aliquots was extracted and chromatographically profiled to establish the nature of the aged residues while the other two 10-gram portions were used for the soil column leaching experiments.
- Preparation of Soil Columns: Columns used in this study were made of plastic and were set upright by a clamp on a support stand. Each column was approximately 520 mm in length and 35 mm in width (diameter). At the bottom of each column was a fine mesh wire screen to support the soil in the column. A funnel at the bottom of the column allowed for the collection of leachate. Both the screen and the funnel were firmly attached to the column with silicone sealant.
The appropriate soil intended for preparing each column was packed by filling each column to the 450-mm mark, tapping with a silicone stopper attached to a piece of PVC pipe and repeating the process until the soil did not settle below the 450-mm mark. This procedure allowed for an even distribution of the soil within the column. The bulk density of each column was then calculated based on the amount of soil added to the column (on a dry weight basis) and the height of the packed column. Following column packing, the columns were saturated with 0.01 N CaCl2 overnight. Heights of the soil columns were determined after allowing the soil in the columns to settle after the saturation period. Void volumes (difference between the amount of CaCl2 solution added to the top of the column and the amount eluted) were also determined prior to conducting the leaching experiment.
The reservoir holding the 0.01 N CaCl2 solution intended for percolation through the soil columns was a 500-mL plastic bottle which was fitted with two silicone stoppers and a glass tube. The larger stopper, with a diameter of 35 mm, fit snugly into the top of the leaching column and allowed for slow delivery of the CaCl2 solution while still maintaining a constant head pressure above the soil in the column.
At the bottom, the leaching columns were connected by silicone tubing to volatile trapping trains comprised of polyurethane foam plugs, ethylene glycol and two 10 % potassium hydroxide solutions for the purpose of on-column trapping of volatiles. Connected to the top of each column was also a volatile trapping solution (10 % potassium hydroxide solution). The headspace of the Erlenmeyer flasks used for leachate collection was purged for at least 15 minutes prior to removal from the column.
- Column Leaching Procedure: To the top of each column was applied a 10-g (dry weight) aliquot of each test material-fortified, aerobically aged soil (at its approximate half-life), topped by a uniform 10-g ( dry weight) layer of untreated soil. A piece of glass wool that allowed for the even distribution of water and minimized physical damage to the soil surface from the applied CaCl2 solution covered each column. Each soil type used for the aged-soil column leaching test was done in duplicate.
Column percolation was initiated by filling the reservoir bottle above each soil column with 490 mL of 0.01 N CaCl2 solution, a volume approximating the 20 inches (50.8 cm) times the cross-sectional area of the column, as specified in the Guideline. Column effluent (leachate) was collected in 6 or 7 individual fractions. Prior to removal of a leachate fraction, the headspace above the collected leachate was purged via vacuum for at least 15 minutes to capture any on-column volatiles. For quantitation, aliquots (2 to 5 mL) of the individual leachate fractions were radioassayed in duplicate by LSC. Leachate fractions containing ~ 5 % of the column-applied radiocarbon were profiled by HPLC-RAM, either directly or after partitioning first with ethyl acetate.
In order to collect at least five individual leachate fractions, slow-percolating soil columns (sandy loam A (soil # 1) and clay) were clamped at the end of a working day to stop percolation and restarted the following morning.
When column percolation was complete, the soil columns were wrapped in aluminum foil and placed in a freezer overnight. Columns were then removed from the freezer and allowed to thaw slightly. The sandy loam A and B soil (# 1 and 2) columns were extruded by using a wooden dowel of a diameter slightly less than the column diameter and wrapped in a piece of Teflon® sheeting. During extrusion, the soil column was cut intact into approximately 6-cm segments with a knife and placed into stainless steel bowls. The clay and sand columns were cut intact into approximately 6-cm segments using a PVC pipe cutter. Individual soil segments were well mixed using stainless steel spatulas and then placed into amber bottles. After determining soil moisture content, aliquots of the soil segments were combusted to quantify radioactive residues present in each soil segment. Individual soil segments containing ≥ 10 % of column-applied radiocarbon were extracted and chromatographically profiled.
- Analysis of Soil Column Segments and Leachate: Aliquots of soil column segments (approximately 50 grams [dry weight]) containing significant radioactive residues (i.e., ≥ 10 % of the column-applied radioactivity) were placed in centrifuge bottles and extracted with 2 x 100 mL of acetonitrile/ water/ glacial acetic acid (80/ 20/ 2.5, v/v/v) by shaking at 250 rpm for 15 minutes. Extracted soil was separated from extraction solvent by centrifugation at 1 000 rpm for 20 minutes. The volume of the combined supernatant (extraction solvent) was recorded and 3 x 1-mL aliquots quantified by LSC. Non-extractable radioactive residues were quantified by combustion analysis.
In soil segments where extractable radioactivity exceeded 10 % of column-applied (uppermost segments from sandy loam B (soil # 2) and sand soils), a 75-mL aliquot of the soil extract was transferred to a round bottom flask and the acetonitrile component of the extraction mixture was removed by rotary evaporation. The remaining acidic aqueous portion was partitioned with three volumes of ethyl acetate. Each ethyl acetate portion was subsequently dried with anhydrous sodium sulfate and combined with the others. After recording the volumes of the combined ethyl acetate and aqueous/acidic phases, 2 x 0.5 mL and 1 x 1 mL aliquots, respectively, were quantified by LSC. For HPLC-RAM profiling, the combined ethyl acetate phase was concentrated to near dryness by rotary evaporation and redissolved in 1 mL of acetonitrile. After quantifying radioactivity (3 x 0.1 mL) by LSC, an aliquot was profiled by HPLC-RAM. Since less than 5 % of initial radioactivity remained in any aqueous/acidic phase, no further analysis of these fractions were undertaken.
Aliquots of leachate fractions generally containing more than 5 % of column-applied radioactivity were adjusted to an approximate pH of 3 with concentrated hydrochloric acid and partitioned with two volumes of ethyl acetate. Each ethyl acetate portion was dried with anhydrous sodium sulfate, combined and its volume recorded. Aliquots (2 x 0.5 mL) were radioassayed by LSC for quantitative purposes. A 1-mL aliquot of the post-partitioned aqueous phase was also radioassayed (LSC). The combined ethyl acetate fraction was concentrated to near dryness under a gentle stream of nitrogen and redissolved in 1 mL of acetonitrile. Radioactivity was quantified using 3 x 0.1-mL aliquots and another aliquot was used for HPLC-RAM profiling. Since radioassay of post-extracted acidic/aqueous phases was below the minimal detection limit for LSC, they were not further analysed.
- HPLC-RAM Analysis
HPLC-RAM profiling of soil extracts from the pilot aerobic soil metabolism study was performed for the purpose of identifying and quantifying remaining test material at various sampling intervals. The HPLC instrumentation consisted of a Waters Model 510 pump, a Hewlett-Packard Model 1050 injector, an Autochrom M-112-2 CIM gradient, a Kratos 757 UV detector and a Radiomatic A280 detector. The UV detector was used to monitor nonradiolabeled reference standards. The conditions for each system are provided below.
HPLC System 1 (employed for days 0, 2 and 6 soil extracts)
Column: Metachem Spherisorb ODS-2 (5 μm, 4.6 mm x 250 mm)
Mobile Phases: Solvent A: 0.1 % trifluoroacetic acid in reagent water; Solvent B: 100 % acetonitrile
Flow Rate: 1.0 mL/minute
Gradient Program: 0 min (60 % A, 40 % B), 30 min (40 % A, 60 % B), 31 min (60 % A, 40 % B)
Equilibrium Delay: 15 min
UV Detection: 220 nm
Run Time: 31 min
Injection Volume: 10 μL (reference standards); 100 μL (samples)
Flow Cell Volume: 1 000 μL
Cocktail Type: Flo-Scint II
Cocktail Flow Rate: 3.0 mL/min
HPLC System 2 (used for day 12 soil extracts)
Column: Metachem Spherisorb ODS-2 (5 μm, 4.6 mm x 250 mm)
Mobile Phases: Solvent A: 0.1 % trifluoroacetic acid in reagent water; Solvent B: 100 % acetonitrile
Flow Rate: 1.0 mL/minute
Gradient Program: 0 min (60 % A, 40 % B), 20 min (60 % A, 40 % B), 30 min (40 % A, 60 % B), 31 min (0 % A, 100 % B), 36 min (0 % A, 100 % B), 37 min (60 % A, 40 % B)
Equilibrium Delay: 15 min
UV Detection: 220 nm
Run Time: 37 min
Injection Volume: 10 μL (reference standards); 50, 100 μL (samples)
Flow Cell Volume: 1 000 μL
Cocktail Type: Flo-Scint II
Cocktail Flow Rate: 3.0 mL/min
HPLC System 3 (used for either direct analysis of leachate fractions or ethyl acetate partitions of leachate fractions)
Column: Metachem Spherisorb ODS-2 (5 μm, 4.6 mm x 250 mm)
Mobile Phases: Solvent A: 0.1 % trifluoroacetic acid in reagent water; Solvent B: 100 % acetonitrile
Flow Rate: 1.0 mL/minute
Gradient Program: 0 min (100 % A, 0% B), 5 min (100 % A, 0 % B), 59 min (0 % A, 100 % B), 60 min (100 % A, 0 % B)
Equilibrium Delay: 15 min
UV Detection: 220 nm
Run Time: 60 min
Injection Volume: 10 μL (reference standards); 50 μL (samples)
Flow Cell Volume: 1 000 μL
Cocktail Type: Flo-Scint II
Cocktail Flow Rate: 3.0 mL/min
HPLC column recoveries for the definitive aged-soil column leaching study had a mean ± standard deviation of 104.9 ± 15.2 %.
- LSC and Oxidiser Instrumentation
Quantitative radioassay by LSC was accomplished with a Beckman LS 3801, LS 1801 or LS 5000 TD Liquid Scintillation Counter. All soil combustions were performed with a Packard Model 306 or 307 Tri-carb Oxidiser. Radioactive 14CO2 formed was trapped in Carbosorb® and flushed into scintillation vials with Perma-Fluor E+® scintillation cocktail and quantified by LSC.
Counting efficiencies of all experimental samples were determined using an external standard and a factory prepared calibration curve. All test samples were counted for a maximum of 5 minutes or until a 2 sigma error of 5 % was attained. Oxidiser efficiency (i.e., instrumental) was determined prior to analysing each set of samples by combusting and counting the radioactivity associated with a standard reference material (Spec-Chec™ [14C] standard) and comparing the measured value to that of a concurrently fortified Spec-Chec™ standard added directly to scintillation cocktail. Since the oxidiser recovery was typically greater than 98 %, experimental data were not corrected. Oxidiser efficiency for the combustion of soil (i.e., efficiency of burning soil) was determined in triplicate at the time of soil combustions by combusting and quantifying a known amount of 14C-test material (30 μL of a 5.27 mg/L solution representing 18 490 dpm [based on a specific activity of 116 952 dpm/μg]) applied to approximately 0.1 to 0.2 g aliquots of control soils. Radioactivity recovered after soil combustion was compared to the same amount of [14C]test material solution added directly to scintillation cocktail and quantified by LSC. Mean efficiencies ranged from 84.5 % to 105.7 %. As a result, quantitation of soil-bound radioactivity was corrected for oxidiser efficiency.
- Liquid Chromatography/Electrospray-Mass Spectrometry (LC/ES-MS)
For confirming test material in selected leachate fractions (from sand and clay column leaching experiments), electrospray LC/MS was conducted using the following conditions and instruments:
Instruments: Perkin-Elmer Sciex APl-100 mass spectrometer equipped with a Heated Nebulizer (HN) Interface. Instrumentation also included a Hewlett-Packard Model 1050 gradient solvent pump and a Hewlett-Packard Model 1050 autosampler.
Instrumental Conditions:
HPLC Parameters:
Column: Metachem Spherisorb C18 (5 μm, 250 x 4.6 mm)
Mobile Phase: Acetonitrile: Water: Acetic Acid (70:30:0.1)
Flow Rate: 1.0 mL/min
Injection Volume: 20 μL
MS Parameters:
Interface: Heated Nebulizer Inlet
HN Temperature: 500 °C
Nebulizer Gas: Nitrogen
Auxiliary Gas: Air
Nebulizer Current: -3.5 μA
Orifice Potential: -15 V
Acquisition Mode: Negative Ion
Scan Range: 210 - 220 amu
- Test Material as Reference Standard: Note that the racemic mixture of the test material was used as the reference standard for HPLC and LC-ES/MS analyses rather than the test material itself (the optical isomer). Since the chromatographic and mass spectral properties of the racemic and the optical isomer are identical under the conditions employed in this study, the racemic serves as a valid representation of the optical isomer (test material).
- Detection Limits: For LSC radioassays, the limit of detection ranged, in general, from 40 to 60 cpm (53 to 80 dpm; counter efficiency = 75 %, typical for combustion of soil samples). Based on an experimentally determined specific activity of 116 952 dpm/μg, the limit of detection was equivalent to 0.0005 to 0.0007 μg of test material (or equivalents).
For HPLC-RAM profiling, the typical instrumental background for the flow-through radiometric detector is 100 dpm. The limit of detection is considered to be 200 dpm, representing twice the typical background. Therefore, an individual, baseline-resolved chromatographic peak would have a limit of instrumental detection of 0.002 μg. Based on typical injection volumes ranging from 100 to 200 μL, the limit of detection would range from 0.009 μg/mL to 0.02 μg/mL (ppm).
- Storage Stability: For the definitive study, HPLC-RAM profiling of soil extracts of aged soils at the approximate half-life of the test material was performed within 10 days of soil extraction for the purpose of characterising the nature of the extractable radioactive residues to be applied to soil columns.
Leachate fractions, generally representing more than 5 % of column-applied radioactivity, were initially analysed directly by HPLC-RAM within 20 days of collection. However, because of the relatively large volume of leachate and the minimal radioactivity present (e.g., 250 dpm/250 μL in a typical injection), subsequent analysis involved partitioning the leachate fraction with ethyl acetate, removing the ethyl acetate under a stream of nitrogen, redissolving the organo-soluble residue in acetonitrile and profiling an aliquot of the leachate extract by HPLC-RAM. These analyses occurred as long as 3 months after initial leachate collections. However, since all subsequent leachate analyses indicated only the presence of test material, no degradation resulting from storage occurred.
Selected soil column segments containing more than 10 % of column-applied radioactivity (sandy loam B and sand) were profiled by HPLC-RAM. The soil column segments from the replicate sandy loam B columns were extracted within 2 weeks of completing the column leaching and initially profiled by HPLC-RAM 9 days later. Because radioactivity was minimal, the soil extracts were later partitioned with ethyl acetate, concentrated and re-profiled by HPLC-RAM approximately 2 months after the initial analysis. The soil column segments from the replicate sand columns were extracted approximately 2½ months after the completion of the leaching procedure. The soil column segment extracts were partitioned with ethyl acetate, concentrated and profiled by HPLC-RAM within 6 days of extraction. Since the test material accounted for at least 90 % of the HPLC column-eluting radioactivity for both replicates, no apparent effect of storage was evident.
Mass spectral confirmation of test material in leachate fractions of sand and clay soil columns was accomplished approximately 7½ months after initial collection. Since the purpose of the analysis was to provide a qualitative confirmation of identity to supplement the original HPLC-RAM identification, storage had no effect on the validity of the procedure.
Soil column segments, soil extracts and leachates were stored either refrigerated or in a freezer prior to chromatographic or mass spectral analysis.
- Column Elution: To simulate a worst-case water-saturated condition, the column was eluted with a volume of water (actually 0.01 N calcium chloride) equal to 20 inches (50.8 cm) times the cross-sectional area of the column. The column diameter was 3.5 cm. The cross-sectional area of the column equals πr^2. Therefore, the required elution volume was:
50.8 cm x πr^2 = 50.8 cm x 3.1416 x (1.75 cm)^2
= 488.8 cm^3
= 488.8 mL
Therefore, after rounding, the volume of 0.01 N calcium chloride needed for elution of each soil leaching column was 490 mL. - Sample No.:
- #1
- Type:
- Kd
- Value:
- 0.438 dimensionless
- pH:
- 7.4
- Temp.:
- 25 °C
- Matrix:
- Soil (sandy loam A)
- % Org. carbon:
- 0.9
- Sample No.:
- #4
- Type:
- Kd
- Value:
- 0.392 dimensionless
- pH:
- 7
- Temp.:
- 25 °C
- Matrix:
- Soil (clay)
- % Org. carbon:
- 1.6
- Transformation products:
- yes
- No.:
- #1
- Validity criteria fulfilled:
- not specified
- Conclusions:
- Column leaching experiments for aged residues of 14C-test material indicated that the residual parent compound was quite mobile in all four soils tested (two different sandy loams, a sand and a clay). Although 4-chloro-2-methylphenol was only minimally extractable (accounting for no more than 1.5 % of the column-applied radioactivity in any soil type), it was not detected in any leachate fractions or any soil column segment below the uppermost segment. Thus, it appears to have little or no soil mobility. The remaining degradates comprising the aged radioactive residues of test material corresponded to soil-bound entities and also showed no potential for soil movement.
Overall, the results indicated that residual test material, not yet degraded by soil microbes, has significant mobility in a variety of soil types. However, test material degradates appear to have little or no mobility because they are strongly soil-associated. Soil half-lives of the test material are relatively short (2.5 to 10.5 days in this study), therefore the potential for the test material to move in the soil column may be balanced by its relatively rapid soil degradation kinetics. - Executive summary:
An aged-soil column leaching study was conducted with a 14C-version of the test material. The study was conducted in accordance with the standardised guideline EPA/FIFRA Subdivision N Series 163-1 Guideline and under GLP conditions.
Prior to column leaching, a pilot [14C]test material soil metabolism study was conducted at an application rate of 2.4 lbs a.i./acre, representing the highest seasonal application rate of test material on turf. Based on chromatographic (HPLC-RAM) analysis of soil extracts and the assumption of first-order kinetics, soil half-lives of test material were calculated to be 7.5, 2.5, 10.5 and 8.2 days in two different sandy loam soils (designated sandy loam A and sandy loam B), a sand, and a clay, respectively, at an incubation temperature of 25 ± 1 °C. (Sandy loam B was initially thought to be a silt loam but was subsequently characterised as a sandy loam soil). In addition to providing half-life information, the pilot soil metabolism study also showed that the test material was significantly metabolised to soil-bound (non-extractable) residues (accounting for 21.0 % [sand] to 44.4 % [sandy loam B] of the applied radioactivity in the four soils after 12 days of incubation). Production of volatiles was also significant during the pilot soil metabolism study ranging from 16.3 % (clay) to 22.9 % (sandy loam B) in the four soils after 12 days of incubation.
Based on the results of the pilot soil metabolism study, [14C]test material (100 % radiopurity, specific activity of 116 952 dpm/μg [11.3 mCi/mmol]) was applied to the four soil types at the highest seasonal application rate and aged in each soil to its approximate half-life. Chromatographic (HPLC-RAM) analysis of soil extracts indicated that the test material accounted for 49.0, 52.1, 63.0 and 74.0 % of the total radioactive residues for sandy loam B, sandy loam A, sand, and clay, respectively, thereby achieving the objective of reaching the approximate half-life. Ten-gram aliquots of aged radioactive residues from each soil type were then applied to corresponding soil columns (in duplicate) with a diameter of 35 mm and heights ranging from 301 to 352 mm after saturation and soil settling. Soil columns were percolated with the equivalent of 20-acre inches of water (0.01 N CaCl2), maintaining a head pressure throughout the leaching experiment. Percolation rates were quite variable depending on soil type and ranged from 7.60 mL/min in a replicate sand soil column to 0.003 mL/min in a replicate clay column. Movement of radioactive residues (test material and its degradates) was monitored and quantified in soil column segments and leachate fractions. Production of volatiles during column leaching (negligible, no more than 0.2 % of column-applied radioactivity for any column) was quantified by trapping in various media.
In all four soil column leaching experiments, a significant percentage of column-applied radioactivity was collected in leachates, accounting for mean values of 54.0, 30.5, 46.6 and 69.5 % in sandy loam A and B, sand, and clay, respectively. In all individual leachate fractions (minimum of six per soil column) generally containing greater than 5 % of the column-applied radioactivity, the only detectable eluting radioactive residue corresponded to the test material. Identification of the test material was based on HPLC-RAM profiling, comparing the retention time of a reference standard to the radioactive residue in leachate samples. Confirmation of identity in selected leachate samples was subsequently accomplished by HPLC/[electrospray]-mass spectrometry.
Radioactive residues not found in leachate fractions were primarily found in the uppermost soil segment of soil columns (ranging from 19.3 % [clay, replicate column B] to 46.0 % [sandy loam B, replicate column B]). These segments represented primarily soil-bound residues since the majority of radioactivity was non-extractable. In instances where extractable radioactivity from these segments was ~ 10 % of column-applied (sandy loam B and sand columns), HPLC-RAM profiling revealed the predominant presence of the test material, with small amounts of 4-chloro-2-methyl phenol, the latter not contributing more than 1.3 % of the column-applied radioactivity.
Mean material balances (summation of radioactivity in soil column segments, leachates and volatile traps compared to column-applied radioactivity; averaged for two replicates/soil type) during column leaching were determined to be 91.2, 88.4, 82.2, and 116 % for sandy loam A and B, sand, and clay, respectively.
The distribution coefficients (Kd) of the test material in the soil columns were calculated, based on the paper of Swoboda and Thomas (1968), to average 0.438 and 0.392 for sandy loam A and clay, respectively. Kd values for sandy loam B and sand could not be calculated since less than 50 % of the column-applied radioactivity was collected in the leachate for either replicate soil column.
Based on the results of this study, the test material was relatively rapidly metabolised in all the soils tested to produce degradates which were either volatile (CO2), extractable but not soil mobile (4-chloro-2-methylphenol, which was only found in uppermost soil column segments), or non-extractable and non-leachable entities (soil-bound residues either tightly associated with or incorporated into humic substances). The sole component of the test material aged residue showing significant soil mobility was unmetabolised test material. The amount of radioactivity in the leachate was proportional to the amount of test material in the aged soil applied to the soil columns. Soil half-lives of the test material were relatively short (2.5 to 10.5 days), therefore the potential for the test material to move in the soil column may be balanced by its relatively rapid soil degradation kinetics.
Referenceopen allclose all
Definitive Phase, Mean Concentration of Test Material in the Solid and Liquid Phases at the End of the Adsorption Period
Concentration (mg L^-1) |
Soil (μg g^-1) |
Solution (μg g^-1) |
% Adsorbed |
Calke |
|||
1 |
0.397 |
0.825 |
19.3 ± 0.4 |
0.4 |
0.224 |
0.278 |
27.7 ± 0.0 |
0.1 |
0.062 |
0.068 |
30.7 ± 0.4 |
0.04 |
0.026 |
0.026 |
32.3 ± 0.4 |
0.01 |
0.007 |
0.007 |
33.7 ± 0.1 |
South Witham |
|||
1 |
0.363 |
0.835 |
17.7 ± 0.1 |
0.4 |
0.174 |
0.313 |
21.6 ± 0.2 |
0.1 |
0.049 |
0.075 |
24.4 ± 0.6 |
0.04 |
0.020 |
0.030 |
25.2 ± 0.1 |
0.01 |
0.006 |
0.008 |
26.8 ± 0.1 |
Lockington |
|||
1 |
0.542 |
0.857 |
17.8 ± 0.1 |
0.4 |
0.253 |
0.325 |
21.1 ± 0.8 |
0.1 |
0.076 |
0.077 |
25.4 ± 0.6 |
0.04 |
0.032 |
0.030 |
26.9 ± 0.7 |
0.01 |
0.010 |
0.008 |
30.6 ± 1.2 |
Hagen |
|||
1 |
0.659 |
0.682 |
32.1 ± 0.3 |
0.4 |
0.289 |
0.255 |
35.8 ± 0.5 |
0.1 |
0.074 |
0.062 |
37.1 ± 0.0 |
0.04 |
0.031 |
0.024 |
38.9 ± 0.5 |
0.01 |
0.009 |
0.006 |
40.6 ± 0.1 |
Definitive Phase, Mean Concentration of Test Material in the Solid and Liquid Phases at the End of the Desorption Period
Concentration (mg L^-1) |
Soil (μg g^-1) |
Solution (μg g^-1) |
% Adsorbed |
Calke |
|||
1 |
0.258 |
0.252 |
34.9 ± 1.9 |
0.4 |
0.148 |
0.094 |
33.7 ± 0.6 |
0.1 |
0.041 |
0.024 |
33.5 ± 0.1 |
0.04 |
0.018 |
0.009 |
31.9 ± 0.8 |
0.01 |
0.005 |
0.003 |
31.0 ± 0.7 |
South Witham |
|||
1 |
0.235 |
0.261 |
35.4 ± 0.7 |
0.4 |
0.118 |
0.101 |
32.1 ± 0.7 |
0.1 |
0.032 |
0.026 |
34.7 ± 2.3 |
0.04 |
0.014 |
0.010 |
33.4 ± 0.8 |
0.01 |
0.004 |
0.003 |
34.7 ± 1.4 |
Lockington |
|||
1 |
0.334 |
0.251 |
38.4 ± 1.1 |
0.4 |
0.159 |
0.098 |
37.1 ± 1.6 |
0.1 |
0.048 |
0.025 |
36.7 ± 2.9 |
0.04 |
0.021 |
0.010 |
33.1 ± 0.9 |
0.01 |
0.007 |
0.003 |
34.2 ± 0.1 |
Hagen |
|||
1 |
0.406 |
0.251 |
38.3 ± 0.4 |
0.4 |
0.186 |
0.098 |
35.6 ± 1.1 |
0.1 |
0.048 |
0.025 |
35.7 ± 0.7 |
0.04 |
0.021 |
0.010 |
33.9 ± 1.0 |
0.01 |
0.006 |
0.003 |
34.1 ± 0.3 |
Expressed as a percentage of the initially adsorbed material; after the desorption cycle
Freundlich Adsorption and Desorption Coefficients and Constants
Soil |
Adsorption |
Desorption |
||||||
Kf (mL g^-1) |
1/n |
R^2 |
Koc (mL g^-1) |
Kdes (mL g^-1) |
1/n |
R^2 |
Kocdes (mL g^-1) |
|
Calke |
0.56 |
0.852 |
0.99 |
18 |
1.00 |
0.869 |
0.99 |
32 |
South Witham |
0.46 |
0.892 |
1.00 |
12 |
0.88 |
0.915 |
1.00 |
24 |
Lockington |
0.64 |
0.853 |
1.00 |
21 |
1.15 |
0.866 |
1.00 |
38 |
Hagen |
0.98 |
0.926 |
1.00 |
34 |
1.55 |
0.936 |
1.00 |
54 |
Mean |
0.66 |
0.881 |
1.00 |
21 |
1.15 |
0.897 |
1.00 |
37 |
Table 1: Texture, pH, content of humus and WHC of the soils and adsorption of the test material described by Kd and Koc.
Depth (cm) |
Clay |
Silt |
Coarse Silt |
Fine Sand |
Coarse Sand |
Humus |
pH (H2O) |
WHC |
Adsorption |
|
Kd |
Koc |
|||||||||
Flakkebjerg soil |
||||||||||
0 - 30 |
15.8 |
18.4 |
|
44.2 |
19.2 |
2.4 |
6.9 |
31 |
0.12 |
8.4 |
Roskilde soil |
||||||||||
0 - 30 |
13.2 |
14.1 |
14.5 |
31.5 |
24.0 |
2.6 |
6.7 |
31 |
0.20 |
13.3 |
Jyndevad soil |
||||||||||
0 – 25 |
4.0 |
3.8 |
2.8 |
20.4 |
66.5 |
2.4 |
6.6 |
29 |
0.07 |
5.3 |
Clay: < 0.002 mm; silt: 0.002 - 0.02 mm; coarse silt: 0.02 - 0.063 mm; fine sand: 0.063 - 0.2 mm; coarse sand: 0.2 - 2 mm; humus: %C x 1.72.
WHC: Water holding capacity, % of wet soil.
Table 1: The Concentration of the Test Material in Solution as Function of Time
Time (h) |
Amount of Test Material |
||
Soil A (μg/mL) |
Soil B (μg/mL) |
Soil C (μg/mL) |
|
0 |
5.00 |
5.0 |
5.00 |
1 |
4.14 |
4.29 |
4.37 |
4 |
4.05 |
4.22 |
4.36 |
20 |
3.98 |
4.03 |
4.21 |
48 |
3.87 |
3.87 |
4.03 |
Table 2: Distribution of the Test Material Between Solution and Soil After 48 Hours
Amount of Test Material (µg) |
Soil A |
Soil B |
Soil C |
||||
Nominal |
Found |
Solution (μg/mL) |
Adsorbed (μg/g) |
Solution (μg/mL) |
Adsorbed (μg/g) |
Solution (μg/mL) |
Adsorbed (μg/g) |
5 |
5.05 |
0.30 |
2.05 |
0.34 |
1.65 |
0.35 |
1.52 |
10 |
10.4 |
0.67 |
3.35 |
0.72 |
2.87 |
0.74 |
2.62 |
20 |
19.78 |
1.38 |
5.92 |
1.50 |
4.78 |
1.50 |
4.74 |
50 |
48.45 |
3.78 |
10.68 |
3.94 |
9.05 |
3.94 |
9.10 |
From these data adsorption isotherms were plotted. The adsorption constants were calculated according to the equation of Freundlich:
Cads= K * Ceq^(1/N)
or
log Cads = log K + 1/N log Ceq
The calculated values for K, 1/N and the coefficient of regression (r) are given below. Adsorption constants were also calculated based on organic matter (Kom).
Table 3: Calculated Values for K, Kom, 1/N and r
Soil |
K (mL/g) |
Kom (mL/g) |
1/N |
r |
A |
4.5 |
80 |
0.66 |
0.99 |
B |
3.5 |
97 |
0.69 |
0.99 |
C |
3.3 |
78 |
0.75 |
0.99 |
Table 4: Recovery of Radioactivity (results of the extraction of the soils used in the adsorption test with 5 mg.L^-1)
Soil |
% Radioactivity in |
||
CaCl2 |
Extract |
Total |
|
A |
78 |
16 |
94 |
B |
81 |
18 |
99 |
C |
81 |
17 |
98 |
Nature of Test Material Aged Radioactive Residues
After incubating [14C]test material-fortified soils to their approximate half-life based on results from the pilot soil metabolism study, 10-gram aliquots were extracted to characterise the nature of the aged residues which would be subsequently applied to the leaching columns. Non-extractable radioactivity (soil-bound residues) was quantified by combustion analysis to completely characterise column-applied radioactivity. HPLC-RAM chromatograms of soil extracts indicated that the test material was, by far, the major extractable radioactive residue. Small quantities (ranging from 0.4 % of the applied dose in sandy loam A to 3.0 % in clay) of 4-chloro-2-methyl phenol were also detected in HPLC-RAM profiles of soil extracts but were quite minor compared to residual parent compound. Other metabolites were either evident as volatiles or as soil-bound residues. Also included is the percentage of test material (% of applied dose) remaining at the approximate half-life.
Residual test material ranged from 49.0 to 63.0 % of the applied dose for sandy loam A, sandy loam B and sand, values quite close to half-life. For clay, the initial calculation from the pilot soil metabolism study indicated a half-life of 5.5 days, based on the assumption of first-order kinetics. However, subsequent recalculation (at the time of reporting) indicated a slightly prolonged half-life of 8.24 days. Since aging of test material in the clay soil was based on the original kinetic calculation (i.e., based on a half-life of 5.5 days), the percentage of applied test material was experimentally determined to be 71.6 %, a value somewhat short of achieving the target of 50 %. Nevertheless, clay soil leaching columns were initiated since, even though half-life had not been reached, a sufficient degradate component of the aged residues was still present to accurately evaluate both the soil mobility of parent compound and significant degradates.
Results of Column Leaching Experiments
- Sandy Loam A (Soil # 1)
The uppermost soil segment of each replicate column (0 to 8 cm) retained 36.2 to 38.0 % of the column-applied radioactivity after all of the 490 mL of 0.01 N CaCl2 had percolated through the columns. Since only 11.4 to 11.6 % of the segment (representing 4.2 to 4.3 % of the column-applied radioactivity) was extractable, HPLC-RAM profiling was not done. Radioactive residues were not detected in column segments below 0 to 8 cm for either replicate column. However, 53.8 to 54.2 % of the column-applied radioactivity was sufficiently mobile to travel through the replicate soil columns and reach the leachate collection flasks. Leachate fractions (ethyl acetate partitions) containing greater than 5 % of the column-applied radioactivity (Fractions #2, #3 and #4 and Fractions #2 and #3 from replicate columns A and B, respectively) were profiled by HPLC-RAM with all eluting radioactivity corresponding to test material.
At the end of the leaching experiment, material balances for the two replicate sandy loam A leaching columns (radioactivity in soil segments, leachate fractions and volatile traps) were 90.6 and 91.8 %, respectively, for replicate columns A and B.
- Sandy Loam B (Soil # 2)
The uppermost soil column segment (0 to 6 cm) for both replicates contained the greatest amount of radioactive residues (43.5 and 46.0 % of the column-applied radioactivity, for column A and B, respectively). Only a small percentage (21.9 to 23.8 %) of the soil-associated radioactivity in the 0 to 6 cm segments was extractable, however, representing 10.1 and 10.3 % of the radioactivity applied to the replicate columns. The majority of the extractable radioactivity did, however, correspond to test material (89.8 to 91 .4 %). A small amount of 4-chloro-2-methyl phenol (0.9 to 1.0 % of the column-applied radioactivity) was also present in both 0 to 6 cm segment replicates. The four other 6-cm soil column segments for each sandy loam B replicate column contained small amounts of radioactivity, each individually accounting for less than 5 % of the column-applied radioactivity based on combustion analysis. Therefore, they were not further characterised by extraction and HPLC-RAM profiling.
Somewhat comparable to results observed with sandy loam A leaching columns, 28.2 to 32.7 % of column-applied radioactivity was sufficiently mobile to travel through the replicate sandy loam B columns and reach the leachate collection flasks. Leachate fractions containing greater than 5 % of column-applied radioactivity (Fractions #3, #4 and #5 [column A] and Fractions #4 and #5 [column B]) were profiled by HPLC-RAM with all eluting radioactivity corresponding to test material.
At the end of the leaching experiment, material balances for sandy loam B leaching columns were 89.7 and 87.0 %, for replicate columns A and B, respectively.
- Sand (Soil # 3)
The uppermost soil segment of each replicate column (0 to 9 cm, replicate column A; 0 to 11 cm, replicate column B) retained 34.7 to 35.8 % of the column-applied radioactivity after all of the 490 mL of 0.01 N CaCl2 had percolated through the columns. For replicate column A, 38.8 % of the soil-associated radioactivity in the uppermost 0 to 9 cm segment was extractable, representing 13.4 % of the radioactivity applied to the column. HPLC-RAM profiling indicated that 88.8 % of the eluting radioactivity corresponded to test material (11.9 % of column-applied radioactivity) and 11.2 % corresponded to 4-chloro-2-methyl phenol (1.5 % of column-applied radioactivity). For replicate column B, 35.2 % of the soil-associated radioactivity in the uppermost 0 to 11 cm soil segment was extractable, representing 12.6 % of the radioactivity applied to the column. HPLC-RAM profiling indicated that 93.5 % of the eluting radioactivity corresponded to test material (11.8 % of column-applied radioactivity) and 6.5 % corresponded to 4-chloro-2-methyl phenol (0.8 % of column-applied radioactivity).
The cumulative percentage of column-applied radioactivity which was collected in six leachate fractions represented 47.4 and 46.5 % for replicate columns A and B, respectively. Leachate fractions containing at least 4.9 % of column-applied radioactivity (Fractions #2 and #3 for column A and Fractions #3 and #4 for column B) were profiled by HPLC-RAM and all eluting radioactivity corresponded to test material.
At the end of the leaching experiment, material balances for the two replicate sand leaching columns (radioactivity in soil column segments, leachate fractions and volatile traps) were 82.1 and 82.3 %, respectively, for replicate columns A and B.
- Clay (Soil # 4)
The majority of the radioactive residues associated with the replicate soil columns was found in the uppermost soil column segment (41.8 % of the column applied radioactivity in the 0 to 11 cm segment for replicate column A; 19.3% in the 0 to 8 cm segment for replicate column B). Lesser amounts of radioactivity were found, however, in every column segment of both replicate columns (ranging from 2.6 to 3.5 % for replicate column A and from 2.1 to 7.8 % for replicate column B).
Only a small percentage (11.6 and 10.4 % for column A and B, respectively) of the soil associated radioactivity in the uppermost soil column segment was extractable, representing 4.9 and 2.0 % of the column-applied radioactivity. As a result, these column segments were not chromatographically profiled.
Leachate collection from both replicate soil columns indicated that the majority of the column-applied radioactivity had been sufficiently mobile in the clay soil. The percentage of the column-applied radioactivity which was found in the leachate represented 72.9 and 66.1 % for replicate column A and B, respectively. Leachate Fractions #2, #3, #4 and #5 from replicate column A, and Fractions #3, #4 and #5 from replicate column 8 were subsequently chromatographically profiled by HPLC indicating that all of the radioactivity in the leachate corresponded to test material.
It should be noted that the relatively large percentage of radioactive residues found in the leachate from both replicate clay soil columns is due to the somewhat shorter aging period for test material in the clay soil relative to the other soils. After a 5-day aging period, 71.6 % of the applied radioactivity corresponded to [14C]test material. (The actual first-order kinetic half-life was calculated to be 8.24 days in the pilot soil metabolism study). Therefore, the larger amount of radioactivity in the leachates from the clay soil columns is due to the larger contribution of test material as a component of the aged radioactive residues.
At the conclusion of the leaching experiment, material balances for the two replicate clay soil leaching columns (radioactivity in soil segments, leachate fractions and volatile traps) were 127 and 106 %, respectively, for replicate columns A and B.
> Confirmation of Test Material in Selected Leachate Fractions
The identity of test material as the predominant radioactive residue in leachate fractions was confirmed in selected samples by electrospray LC/MS. Identity was initially established by HPLC-RAM, based on chromatographic retention time comparison with the racemic mixture of test material.
Note that the racemic mixture was used since it would have an identical chromatographic retention time and mass spectrum as the test material when chromatographed on a non-chiral HPLC column.
> Calculation of Kd Values
Kd values were calculated for the test material based on the following equation provided in the paper of Swoboda and Thomas (1968).
Kd = [(VpVv) - 1] Vv/W
Where:
Vp = Volume of effluent to leach one-half of the solute through the column
Vv = Void volume of the column
W = Weight of the soil in the column
Since the radioactive residues detected in all column leachate fractions corresponded exclusively to the test material, Kd values could be determined in all leaching experiments where at least 50 % of the column-applied radioactivity had eluted. Thus, Kd values for the test material in sandy loam A and clay were determined to be 0.438 and 0.392, respectively, based on the mean of duplicate column data. These values indicate the relatively significant mobility of test material in these soil types. Since less than 50 % of the column-applied radioactivity was eluted from the replicate sandy loam B and the sand columns, Kd values for the test material could not be calculated for these soil types.
Nature of Radioactive Residues in Aged-Soil Applied to Soil Columns (% of Applied Radioactivity Unless Otherwise Indicated)
Sandy Loam A |
Sandy Loam B |
Clay |
Sand |
|
Aging period (days) |
6 |
2 |
5 |
11 |
Extractable radioactivity |
52.5 |
51.4 |
74.6 |
66.0 |
Soil-bound radioactivity |
24.7 |
35.6 |
23.5 |
16.2 |
Volatiles† |
10.2 |
5.10 |
3.70 |
12.2 |
Total |
87.4 |
92.1 |
102 |
94.4 |
Test material (% of extractables) |
99.2 |
95.4 |
96.0 |
95.5 |
Test material |
52.1 |
49.0 |
71.6 |
63.0 |
4-chloro-2-methylphenol (% of extractables) |
0.76 |
4.60 |
4.02 |
2.52 |
4-chloro-2-methylphenol |
0.399 |
2.36 |
3.00 |
1.66 |
Based on HPLC-RAM quantitation.
† For all soil types, at least 99 % of the trapped volatile radioactivity produced during the aging period was found in the potassium hydroxide solutions, putatively indicating 14CO2.
Description of key information
Adsorption/Desorption: Simmonds (2010)
Under the conditions of this study, the mean determined Koc and Koc des values were 21 and 37 mL g^-1 respectively, with corresponding mean 1/n values of 0.881 and 0.900. Thus, according to Briggs, the test material can be classified as being mobile in soil and, according to the McCall classification, as having very high mobility in soil.
Adsorption Supporting Study: Hartley (1996)
Column leaching experiments for aged residues of 14C-test material indicated that the residual parent compound was quite mobile in all four soils tested (two different sandy loams, a sand and a clay). Although 4-chloro-2-methylphenol was only minimally extractable (accounting for no more than 1.5 % of the column-applied radioactivity in any soil type), it was not detected in any leachate fractions or any soil column segment below the uppermost segment. Thus, it appears to have little or no soil mobility. The remaining degradates comprising the aged radioactive residues of test material corresponded to soil-bound entities and also showed no potential for soil movement.
Overall, the results indicated that residual test material, not yet degraded by soil microbes, has significant mobility in a variety of soil types. However, test material degradates appear to have little or no mobility because they are strongly soil-associated. Soil half-lives of the test material are relatively short (2.5 to 10.5 days in this study), therefore the potential for the test material to move in the soil column may be balanced by its relatively rapid soil degradation kinetics.
Adsorption Supporting Study: Helweg (1993)
The adsorption of the test material was low with Kd values of 0.07 - 0.2 and Koc values of 8.4, 13.3 and 5.3 in the three soil types.
Adsorption Supporting Study: Matla & Vonk (1993)
Under the conditions of the study adsorption constant values for the test material based on total soil were 4.5, 3.5 and 3.3 mL.g^-1 for sandy soils A, B and C, respectively. Adsorption constants calculated on organic matter base were 80, 97 and 78 mL.g^-1 for sandy soils A, B and C, respectively.
Key value for chemical safety assessment
- Koc at 20 °C:
- 21
Additional information
Adsorption/Desorption: Simmonds (2010)
The adsorption and desorption characteristics of the test material were studied in four soils using the batch equilibrium method in accordance with the standardised guidelines OECD 106 and US EPA OPPTS 835.1230 under GLP conditions. The study was awarded a reliability score of 1 in accordance with the criteria set forth by Klimisch et al. (1997).
The soils used were Calke sandy loam (UK, pH 5.6, 3.13 % organic carbon), South Witham clay loam (UK, pH 7.2, 3.71 % organic carbon), Lockington sandy clay loam (UK, pH 5.6, 3.07 % organic carbon) and Hagen (Refesol 04-A) loamy sand (Germany, pH 5.5, 2.90 % organic carbon). Due to significant breakdown of the test material observed in the soil: solution ratio preliminary test, subsequent tests were performed using soils that had been sterilised by gamma irradiation. An additional test was carried out during the definitive phase using the South Witham soil, which was treated without pre-equilibration with calcium chloride. As this test showed no significant differences in adsorption, it was excluded from all mean data.
The adsorption phase of the study was carried out using sterile, pre-equilibrated soils with [14C]-test material at concentrations of approximately 1.0, 0.4, 0.1, 0.04 and 0.01 mg L^-1 in the dark and at 20 ± 1 °C for 72 hours for all soils. The equilibration solution used was 0.01 M aqueous CaCl2, with a soil: solution ratio of 1:2 for the Calke sandy loam, the South Witham clay loam and for the Hagen (Refesol 04-A) loamy sand and a 1:3 ratio for the Lockington sandy clay loam. The desorption phase of the study was carried out for 2 hours with fresh 0.01 M aqueous CaCl2 applied to pre-adsorbed soil for one desorption cycle.
The aqueous supernatant after adsorption and desorption was separated by centrifugation and analysed by LSC. After desorption, the soil was extracted with acetonitrile and following extraction, the soil was combusted and the trapped CO2 analysed by LSC. The adsorption parameters were calculated using the Freundlich isotherm.
It was proven during the preliminary tests that greater than 90 % of applied radioactivity could be extracted from the soil after 48 hours adsorption and close to 90 % extractable after 72 hours. In both the preliminary and definitive tests, [14C]-test material accounted for greater than 97 % of the region of interest in all supernatants and extracts analysed by HPLC. It was concluded therefore that no significant degradation of [14C]-test material occurred over the duration of the study.
The overall material balances in the definitive phase were determined by LSC of the supernatants after adsorption, desorption and solvent extraction and combustion of the remaining soils. The overall material balances were 96.8 % for the Calke sandy loam, 98.3 % for the South Witham clay loam, 99.9 % for the Lockington sandy clay loam and 97.0 % for the Hagen (Refesol 04-A) loamy sand.
In the definitive adsorption test, the amount of applied test material adsorbed ranged from 19.3 to 33.7 % in the Calke sandy loam, 17.7 to 26.8 % in the South Witham clay loam, 17.8 to 30.6 % in the Lockington sandy clay loam and 32.1 to 40.6 % in the Hagen (Refesol 04-A) loamy sand.
The calculated adsorption constants (Kf) of the Freundlich isotherms for the soils ranged from 0.46 mL g^-1 in the South Witham clay loam to 0.98 mL g^-1 in the Hagen (Refesol 04-A) loamy sand. The Freundlich exponents (1/n) ranged from 0.852 in the Calke sandy loam to 0.926 in the Hagen (Refesol 04-A) loamy sand, indicating that the concentration of the test material does have a significant effect on the adsorption behaviour in the examined concentration range.
The adsorption Koc values ranged from 12 mL g^-1 in the South Witham clay loam to 34 mL g^-1 in the Hagen (Refesol 04-A) loamy sand.
At the end of the desorption phase, the amount of test material desorbed, expressed as a percentage of the initial amount adsorbed, ranged from 31.0 to 34.9 % in the Calke sandy loam, 32.1 to 35.4 % in the South Witham clay loam, 33.4 to 38.4 % in the Lockington sandy clay loam and 33.9 to 38.3 % in the Hagen (Refesol 04-A) loamy sand.
The desorption Kdes values ranged from 0.88 mL g^-1 in the South Witham clay loam to 1.55 mL g^-1 in the Hagen (Refesol 04-A) loamy sand.
For all soils the data fit to a linear equation was good for both adsorption and desorption with correlation coefficients of 0.99 to 1.00.
Under the conditions of this study, the mean determined Koc and Koc des values were 21 and 37 mL g^-1 respectively, with corresponding mean 1/n values of 0.881 and 0.900. Thus, according to Briggs, the test material can be classified as being mobile in soil and, according to the McCall classification, as having very high mobility in soil.
Adsorption Supporting Study: Hartley (1996)
An aged-soil column leaching study was conducted with a 14C-version of the test material. The study was conducted in accordance with the standardised guideline EPA/FIFRA Subdivision N Series 163-1 Guideline and under GLP conditions.The study was awarded a reliability score of 1 in accordance with the criteria set forth by Klimisch et al. (1997).
Prior to column leaching, a pilot [14C]test material soil metabolism study was conducted at an application rate of 2.4 lbs a.i./acre, representing the highest seasonal application rate of test material on turf. Based on chromatographic (HPLC-RAM) analysis of soil extracts and the assumption of first-order kinetics, soil half-lives of test material were calculated to be 7.5, 2.5, 10.5 and 8.2 days in two different sandy loam soils (designated sandy loam A and sandy loam B), a sand, and a clay, respectively, at an incubation temperature of 25 ± 1 °C. (Sandy loam B was initially thought to be a silt loam but was subsequently characterised as a sandy loam soil). In addition to providing half-life information, the pilot soil metabolism study also showed that the test material was significantly metabolised to soil-bound (non-extractable) residues (accounting for 21.0 % [sand] to 44.4 % [sandy loam B] of the applied radioactivity in the four soils after 12 days of incubation). Production of volatiles was also significant during the pilot soil metabolism study ranging from 16.3 % (clay) to 22.9 % (sandy loam B) in the four soils after 12 days of incubation.
Based on the results of the pilot soil metabolism study, [14C]test material (100 % radiopurity, specific activity of 116 952 dpm/μg [11.3 mCi/mmol]) was applied to the four soil types at the highest seasonal application rate and aged in each soil to its approximate half-life. Chromatographic (HPLC-RAM) analysis of soil extracts indicated that the test material accounted for 49.0, 52.1, 63.0 and 74.0 % of the total radioactive residues for sandy loam B, sandy loam A, sand, and clay, respectively, thereby achieving the objective of reaching the approximate half-life. Ten-gram aliquots of aged radioactive residues from each soil type were then applied to corresponding soil columns (in duplicate) with a diameter of 35 mm and heights ranging from 301 to 352 mm after saturation and soil settling. Soil columns were percolated with the equivalent of 20-acre inches of water (0.01 N CaCl2), maintaining a head pressure throughout the leaching experiment. Percolation rates were quite variable depending on soil type and ranged from 7.60 mL/min in a replicate sand soil column to 0.003 mL/min in a replicate clay column. Movement of radioactive residues (test material and its degradates) was monitored and quantified in soil column segments and leachate fractions. Production of volatiles during column leaching (negligible, no more than 0.2 % of column-applied radioactivity for any column) was quantified by trapping in various media.
In all four soil column leaching experiments, a significant percentage of column-applied radioactivity was collected in leachates, accounting for mean values of 54.0, 30.5, 46.6 and 69.5 % in sandy loam A and B, sand, and clay, respectively. In all individual leachate fractions (minimum of six per soil column) generally containing greater than 5 % of the column-applied radioactivity, the only detectable eluting radioactive residue corresponded to the test material. Identification of the test material was based on HPLC-RAM profiling, comparing the retention time of a reference standard to the radioactive residue in leachate samples. Confirmation of identity in selected leachate samples was subsequently accomplished by HPLC/[electrospray]-mass spectrometry.
Radioactive residues not found in leachate fractions were primarily found in the uppermost soil segment of soil columns (ranging from 19.3 % [clay, replicate column B] to 46.0 % [sandy loam B, replicate column B]). These segments represented primarily soil-bound residues since the majority of radioactivity was non-extractable. In instances where extractable radioactivity from these segments was ~ 10 % of column-applied (sandy loam B and sand columns), HPLC-RAM profiling revealed the predominant presence of the test material, with small amounts of 4-chloro-2-methyl phenol, the latter not contributing more than 1.3 % of the column-applied radioactivity.
Mean material balances (summation of radioactivity in soil column segments, leachates and volatile traps compared to column-applied radioactivity; averaged for two replicates/soil type) during column leaching were determined to be 91.2, 88.4, 82.2, and 116 % for sandy loam A and B, sand, and clay, respectively.
The distribution coefficients (Kd) of the test material in the soil columns were calculated, based on the paper of Swoboda and Thomas (1968), to average 0.438 and 0.392 for sandy loam A and clay, respectively. Kd values for sandy loam B and sand could not be calculated since less than 50 % of the column-applied radioactivity was collected in the leachate for either replicate soil column.
Based on the results of this study, the test material was relatively rapidly metabolised in all the soils tested to produce degradates which were either volatile (CO2), extractable but not soil mobile (4-chloro-2-methylphenol, which was only found in uppermost soil column segments), or non-extractable and non-leachable entities (soil-bound residues either tightly associated with or incorporated into humic substances). The sole component of the test material aged residue showing significant soil mobility was unmetabolised test material. The amount of radioactivity in the leachate was proportional to the amount of test material in the aged soil applied to the soil columns. Soil half-lives of the test material were relatively short (2.5 to 10.5 days), therefore the potential for the test material to move in the soil column may be balanced by its relatively rapid soil degradation kinetics.
Adsorption Supporting Study: Helweg (1993)
Adsorption of 14C-ring-labelled test material was determined according to the OECD Test Guideline Adsorption - Desorption in three different soil types. The study was awarded a reliability score of 2 in accordance with the criteria set forth by Klimisch et al. (1997).
Soils were from three Danish Experimental Farms (#1 Flakkebjerg, #2 Roskilde and #3 Jyndevad, providing sandy loam, sandy loam and coarse sandy soil, respectively).
Adsorption (Kd) of the test material was determined at 20 °C by shaking 5 g of soil with 25 mL 0.01 M CaCl2 added to 14C-test material (5 mg L^-1) for 16 h. To avoid degradation the soils were sterilised by irradiation. The soil was precipitated by centrifugation and 14C was counted in aliquots of the supernatant to calculate concentrations of the test material in soil and water. Koc was calculated from Kd and the organic C contents of the soils.
The adsorption of the test material was low with Kd values of 0.07 - 0.2 and Koc values of 8.4, 13.3 and 5.3 in the three soil types (Flakkebjerg, Roskilde and Jyndevad, respectively). With such low adsorption values the test material is likely to leach if not decomposed in the surface soil.
Adsorption Supporting Study: Matla & Vonk (1993)
The adsorption of the test material to three types of soil was determined essentially according the OECD Guideline 106 and according to the requirements of the Dutch Guidelines and in compliance with GLP. The study was awarded a reliability score of 2 in accordance with the criteria set forth by Klimisch et al. (1997).
The study was carried out to investigate the adsorption of the test material to soil particles of three sandy soil types with a pH (KCl) between 4 and 5. The determinations were carried out by the 'slurry method'. To determine the adsorption constants related to soil organic matter contents (Kom) from the adsorption isotherms, four concentrations of the test material were shaken with soil. The balance of the radioactivity was also determined.
Under the conditions of the study adsorption constant values for the test material based on total soil were 4.5, 3.5 and 3.3 mL.g^-1 for sandy soils A, B and C, respectively. Adsorption constants calculated on organic matter base were 80, 97 and 78 mL.g^-1 for sandy soils A, B and C, respectively.
Desorption studies were not carried out because the adsorption of the test material at the 5 mg.L^-1 level was less than 25 %.
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