Benzene, ethenyl-, ar-bromo derivs.

Adsorption / desorption

Link to relevant study record(s)

Reference

Endpoint:

adsorption / desorption: screening

Type of information:

experimental study

Adequacy of study:

key study

Study period:

from August 28, 2003 to November 13, 2003

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

other: see 'Remark'

Remarks:

Study conducted according to GLP standards. Study conducted according to OECD 121 with the following notable exception: OECD 121 states "For ionisable substances, two tests should be performed with both ionised and non-ionised forms in appropriate buffer solutions but only in cases where at least 10 % of the test compound will be dissociated within pH 5.5 to 7.5." Given that the pKa of the test compound is 6.72 (endpoint study 4.2.1), more than 10% will be dissociated within this pH range, and therefore the two tests should have been conducted.

Qualifier:

according to guideline

Guideline:

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

Deviations:

yes

Remarks:

Details not specified

Principles of method if other than guideline:

or ionisable substances, two tests should be performed with both ionised and non-ionised forms in appropriate buffer solutions but only in cases where at least 10 % of the test compound will be dissociated within pH 5.5 to 7.5. Given that the pKa of the test compound is 6.72 (endpoint study 4.2.1), more than 10% will be dissociated within this pH range, and therefore the two tests should have been conducted.

GLP compliance:

yes

Type of method:

HPLC estimation method

Media:

soil/sewage sludge

Radiolabelling:

no

Test temperature:

Not applicable, the HPLC estimate method was performed; the column temperature was maintained at 40 °C

Details on study design: HPLC method:

PREPARATION OF TEST SUBSTANCE
A stock solution of DBS was prepared by accurately weighing 0.4996 g of the test substance into a 100-mL volumetric flask and bringing to volume using methanol. This primary stock solution contained 5.00 mg/mL of DBS. Subsequent dilution (in duplicate) of the methanol stock solution with the 55:45 (v/v) methanol:water mobile phase solution yielded 50.0-μg DBS/mL secondary solutions. Each injection (100 μL) of these solutions gave a detector response of at least 1200 milli-absorbance units for the peak height of the most sensitive DBS peak.

EQUIPMENT
- Apparatus: Hewlett-Packard Model 1090 High Performance Liquid Chromatograph (HPLC).
- Type, material and dimension of analytical (guard) column: Zorbax CN Column (150 mm x 4.6 mm, 5- µm particle size)
- Detection system: Hewlett-Packard Model 1100 Variable Wavelength (UV) Detector

MOBILE PHASES
- Type: Methanol:HPLC-grade reagent water; 55:45, v/v
- Solutes for dissolving test and reference substances: Methanol (HPLC grade or equivalent.)

DETERMINATION OF DEAD TIME
- Method: By inert substances which are not retained by the column. Prior to the
test substance analyses, the dead time was determined by injecting thiourea.

REFERENCE SUBSTANCES
- Identity: Thiourea (purity 99+%), Acetanilide (purity 99.91%), Phenol (purity 99.99%), Naphthalene (purity 99.9%), Phenanthrene (purity 100%), Alpha-endosulfan (purity 99.5%) and 1,1-bis (p-chlorophenyl)-2,2,2-trichloroethane (DDT) (purity 99.3%).Calibration stock
solutions of the individual reference standards were prepared separately by dissolving each in methanol. Calibration standard solutions were prepared from these stock solutions of the reference standards by diluting appropriately in 55% acetonitrile: 45% HPLC grade reagent water (v/v) mobile phase. The concentrations of the reference standards were adjusted, as necessary, to give a detector response of at least 300 milli-absorbance units for peak height.


DETERMINATION OF RETENTION TIMES
- Quantity of test substance introduced in the column: 100 µL
- Quantity of reference substances: 100 µL
- Intervals of calibration: Reference standard solutions (100 μL injection volume) were injected individually prior to and after the test substance to determine retention times. The test substance solutions were injected following the initial reference standard injections and were analyzed under the same chromatographic conditions

REPETITIONS
- Number of determinations: Performed in duplicate

EVALUATION
- Calculation of capacity factors k': Data Analysis
The capacity factor (k′), was calculated for the each of the test substance chromatographic components and each reference standard using the following equation:
k′ = (tR – t0)/t0
where tR was the retention time of the test substance or reference standard and t0 was the column dead time established with thiourea. A correlation graph of log k′ versus log KOC for the reference standards was plotted and fitted to a regression equation in the form y = mx + b. The calibration curve is presented in Figure 1 in the attached illustration..
- Calculation of retention times: Reference standard solutions (100 µL injection volume) were injected individually prior to and after the test substance to determine retention times.
- Determination of the log Koc value: Log KOC for the test substance was calculated by substituting the calculated logarithm of the capacity factor for DBS into the linear regression equation for the calibration curve

Type:

log Koc

Value:

3.79

Details on results (HPLC method):

Along with analyses of reference standards, a standard of thiourea was injected in duplicate (once near the beginning and once near the end of the HPLC sequence) to determine the analytical column dead time (t0) for use in calculating capacity factors (k′) of the reference standards and the test substance. The mean retention time of the duplicate thiourea standard injections was 2.110 minutes.

Six additional reference standards were analyzed with the test substance. The reference standard series was injected at the beginning and at the end of the HPLC sequence. The capacity factors of each standard were calculated based upon their retention times. Calculated log k′ values for acetanilide, phenol, naphthalene, phenanthrene, alpha-endosulfan and DDT reference standards are presented in Table 3.

Under the chromatographic conditions specified in OECD Guideline 121, DBS eluted as three chromatographically resolved peaks. The first peak, representing the monobromostyrene minor constituent, had a mean peak retention time of 5.435 minutes. The second peak, representing the dibromostyrene major constituent, had a mean peak retention time of 7.067 minutes. The third peak, representing the tribromostyrene minor constituent, had a mean peak retention time of 9.275 minutes. The retention times for all three DBS peaks were bracketed by the reference standards. The corresponding mean log KOC values for DBS were calculated as 3.27 ± 0.000274, 3.79 ± 0.000551 and 4.26 ± 0.000762, respectively, for the three constituents. The calculated mean retention times, capacity factors, and mean log KOC values for DBS are presented in Table 4.

Table 3

Reference Standards Retention Times, Capacity Factors, and log KOC Values

Refernce Standard	Concentration (µg/mL)	Retention Time (minutes)	Capacity Factor (k’)^1,2	log k’^2	Log KOC^3
Acetanilide	50.0	2.725 2.728	0.291 0.293	-0.535 -0.533	1.25
Phenol	50.0	2.763 2.760	0.309 0.308	-0.509 -0.511	1.32
Naphthalene	12.5	5.004 5.005	1.37 1.37	0.137 0.137	2.75
Phenanthrene	25.0	8.657 8.637	3.10 3.09	0.492 0.490	4.09
Alfa-Endosulfan	200	9.114 9.057	3.32 3.29	0.521 0.518	4.09
DDT	100	19.460 19.312	8.22 8.15	0.915 0.911	5.63
1 Capacity factor (k′) is the standard retention time minus the mean column dead time (2.110 minutes for thiourea) divided by the mean column dead time. 2 Values obtained utilizing Excel 2000 in full precision mode. Manual calculations may differ slightly. 3 log KOC values are based on literature values

 

 

Table 4

log  k_OCfor DBS

Sample Number (298C-132-)	Retention Time (minutes)	Mean (x) Standard Deviation (SD)	Capacity Factor (k')^1,2	log k'^2	log kOC^2	Mean (x) Standard Deviation (SD)^2
1 (peak 1)	5.434	x = 5.435	1.58	0.197	3.27	x = 3.27
2 (peak 1)	5.435	SD = 0.000707	1.58	0.198	3.27	SD = 0.000274
1 (peak 2)	7.065	x = 7.067	2.35	0.371	3.79	x = 3.79
2 (peak 2)	7.068	SD = 0.00212	2.35	0.371	3.79	SD = 0.000551
1 (peak 3)	9.272	x = 9.275	3.39	0.531	4.26	x = 4.26
2 (peak 3)	9.278	SD = 0.00424	3.40	0.531	4.26	SD = 0.000762
^1Capacity factor is the sample retention time minus the mean column dead time (2.110 minutes for thiourea) divided by the mean column dead time. ^2Values obtained utilizing Excel 2000 in full precision mode. Manual calculations may differ slightly.

Validity criteria fulfilled:

yes

Remarks:

OECD 121 (Repeatability) states that "Determinations should be run at least in duplicate. The values of log Koc derived from individual measurements should be within a range of 0.25 log unit."

Conclusions:

Under the chromatographic conditions specified in OECD Guideline 121, DBS eluted as three chromatographically resolved peaks. The first peak, representing the monobromostyrene minor constituent, had a mean peak retention time of 5.435 minutes. The second peak, representing the dibromostyrene major constituent, had a mean peak retention time of 7.067 minutes. The third peak, representing the tribromostyrene minor constituent, had a mean peak retention time of 9.275 minutes. The retention times for all three DBS peaks were bracketed by the reference standards. The corresponding mean log KOC values for DBS were calculated as 3.27 ± 0.000274, 3.79 ± 0.000551 and 4.26 ± 0.000762, respectively, for the three constituents.

Executive summary:

In a GLP compliant study conducted according to OECD Guideline 121, the Koc of the test material was determined under chromatographic conditions. DBS eluted as three chromatographically resolved peaks. The first peak, representing the monobromostyrene minor constituent, had a mean peak retention time of 5.435 minutes. The second peak, representing the dibromostyrene major constituent, had a mean peak retention time of 7.067 minutes. The third peak, representing the tribromostyrene minor constituent, had a mean peak retention time of 9.275 minutes. The retention times for all three DBS peaks were bracketed by the reference standards. The corresponding mean log KOC values for DBS were calculated as 3.27 ± 0.000274, 3.79 ± 0.000551 and 4.26 ± 0.000762, respectively, for the three constituents.

Description of key information

Under the chromatographic conditions specified in OECD Guideline 121, DBS eluted as three chromatographically resolved peaks. The first peak, representing the monobromostyrene minor constituent, had a mean peak retention time of 5.435 minutes. The second peak, representing the dibromostyrene major constituent, had a mean peak retention time of 7.067 minutes. The third peak, representing the tribromostyrene minor constituent, had a mean peak retention time of 9.275 minutes. The retention times for all three DBS peaks were bracketed by the reference standards. The corresponding mean log KOC values for DBS were calculated as 3.27 ± 0.000274, 3.79 ± 0.000551 and 4.26 ± 0.000762, respectively, for the three constituents.

Key value for chemical safety assessment

Koc at 20 °C:

3.79

Additional information

The key study Raymond L. Van Hoven, Willard B. Nixon (2003) was performed in compliance with GLP and conducted to the guideline OECD 121. Accordingly the study was assigned a relaibility score of 2 and considered adequate for fulfilment of the data requirement as a stand alone study.