TEST SUBSTANCE: )

DETERMIGOOD LABORATORY PRACTICE COMPLIANCE STATEMENT

SPONSOR:

### QUALITY ASSURANCE STATEMENT

This study was examined for compliance with Good Laboratory Practice Standards as published by the U.S. Environmental Protection Agency in 40 CFR Part 160 andlor 792, 17 August 1989 and OECD Principles of Good Laboratory Practice, (ENVIMCICHEM (98) 17). The dates of all inspections and audits and the dates that any findings were reported to the Study Director and Laboratory Management were as follows:

ACTIVITY: DATE REPORTED TO: DATE CONDUCTED: STUDY DIRECTOR: MANAGEMENT:

Protocol June 7,2005 June 7,2005 September 14,2005

Sample Collection and Processing August 29,2005 August 29,2005 September 7,2005

Data and Draft Report September 12 and 13,2005 September 13,2005 September 15,2005

Final Report September 20,2005 September 20,2005 September 20,2005

All inspections were study-based unless otherwise noted.

Marshall T. Hynson DATE 

# ' L ~ualig~ssurkce Program supe&isor-4-



### REPORT APPROVAL

MANAGEMENT:TABLE OF CONTENTS

Title/Cover Page ............................................................................................................... ...........................1

Good Laboratory Practice Compliance Statement.................................................................................. .....2

Quality Assurance Statement.................................................................................................... ...................3

Report Approval................................................................................................................ ...........................4

Table of Contents.............................................................................................................. ...........................5

Summary........................................................................................................................ ..............................7

Introduc tion................................................................................................................... ...............................8

Objective...................................................................................................................... ................................8

Experimental Design............................................................................................................ ........................8

Materials an d Methods.......................................................................................................... .......................8 Test Substance ................................................................................................................. ....................9 Solvents a nd Reagents .......................................................................................................... ...............9 Test Procedures................................................................................................................ ....................9 Prelimin ary Study ............................................................................................................. ..............9 Definitive Study.............................................................................................................. ................9 Stock and Calibration St andards Preparation .................................................................................... ..9 Analytical Method .............................................................................................................. ...............10 Calibration Curve.............................................................................................................. .................10

Results and Discussion ......................................................................................................... .....................11

Conclusions.................................................................................................................... ............................11

References..................................................................................................................... .............................12TABLE OF CONTENTS - Continued -

TABLES

Table 1 Typical HPLC Operat ional Para meters .....................................................................................13

Table 2 Solubility Results of the Test Subs tance in HPLC-Grade Reagent Water ................................14

FIGURES

Figure 1 Analytical method flowchart for the analysis of the test substance in HPLC-grade reagent water .................................................................................................................. ........15

Figure 2 A representative calibration curv e for the test substance .......................................................16

Figure 3 A representative chromatogram of a low-level (5.00 mg/L) test substance calibration standard ......................................................................................................... .......17

Figure 4 A representative chromatogram of a high-level (50.0 mg/L) test substance calibration standard ......................................................................................................... .......18

Figure 5 A representative chromatogram of a Day 1 water solubility sample 19

APPENDICES

Appendix 1 Protocol, Protocol Ame ndments and Deviation................................................................20

Appendix 2 Personnel Involv ed in the Study .......................................................................................34INTRODUCTION
Tests were conducted to determine the solubility of th e test substance in HPLC-grade reagent water.
This study was conducted for
The analyses of the sa mples were performed using high performance liquid
chromatography with ultraviolet detection (HPLC/ UV). The preliminary portion of this study was
conducted on August 22 and 23, 2005. The definitive portion of this study was conducted between
August 26 and August 29, 2005. The original raw data ge nerated by and a copy
of the final report are filed under Project Numb er 523C-104 in archives located on the

OBJECTIVE
The objective of this study was to determine th e solubility of the test substance at ambient

temperature.

EXPERIMENTAL DESIGN
The water solubility of the test substance was de termined at ambient temperature (approximately
22°C). The test consisted of equilibration of an exce ss amount of test substance with HPLC-grade reagent
water at approximately 22 ° C. Quantitation of soluble test substance in the aqueous phase was performed on
Days 1, 2 and 3.

MATERIALS AND METHODS
This study was conducted according to the proce dures outlined in the corresponding protocol
(Appendix 1). Due to physical properties of the test substance, the test procedure was modified and
performed, based in part, on procedures in the U. S. EPA Product Properties Test Guidelines, OPPTS,
Series: 830.7840, Water Solubility: Column Elution Method; Shake Flask Method (1) utilizing shake flask
methodology. Additional guideline was provided in OECD Guideline for Testing of Chemicals, 105:
Water Solubility (2), and TSCA Title 40 of the Federal C ode of Regulations, Part 796, Section 1840: Water
Solubility (3).- 10 -

standards in 50% methanol : 50% H 2 O. The following shows the diluti on scheme for the set of calibration

standards:

Stock Concentration (mg/mL) Aliquot ( μ L) Final Volume

(mL) Standard Concentration (mg/L)

1.00 1.00 1.00 1.00 1.00 500 1500 2500 3500 5000 100 100 100 100 100 5.00 15.0 25.0 35.0 50.0

Analytical Method
The method used for the analysis of the test s ubstance in HPLC-grade reagent water was based upon
methodology developed by The analytical method consisted of diluting the
samples in 50% methanol : 50% HPLC-grade bottled water and submitting for analysis using high
performance liquid chromatography.

Concentrations of the test substance in the samp les were determined by HPLC using an Agilent
Model 1100 High Performance Liquid Chromatograph (HPLC) equipped with an Agilent Series 1100
Variable Wavelength Detector. Chromatographic separations were achieved using a YMC Pack ODS AM
column (150 mm x 4.6 mm, 3- μ m particle size). Typical instrumental parameters are presented in Table 1
and a method flowchart is provided in Figure 1.

Calibration Curve

Five calibration standards of the test substa nce were prepared in 50% methanol : 50% H 2 O. The
calibration standards, ranging in concentration from 5.00 to 50.0 mg/L, were analyzed concurrently with the
solubility samples. The calibration set was injected at the beginning and end of each analytical sequence.
Linear regression equations were generated using th e sum of the peak area responses of the calibration

standards. The concentration of the test substance in the samples was determined by substituting the sum of
the peak area responses of the samples into the a pplicable linear regression equation. A representativecalibration curve for the test substance is presented in Figure 2. Representative chromatograms of low- and
high-level test substance calibration standards ar e presented in Figures 3 and 4, respectively.

RESULTS AND DISCUSSION
The solubility of the test substance in water at 8500 mg/L was determined on Days 1, 2 and 3. Mean
analytical results were 8098 mg/L ± 31.6 (RSD = 0.390%), 8225 mg/L ± 118 (RSD = 1.43%) and
8261 mg/L ± 134 (RSD = 1.62%) mg/L, respectively. Results are summarized in Table 2. A representative
chromatogram of a Day 1 water solubility sample is presented in Figure 5.

CONCLUSIONS
The solubility of the test substance was determin ed in HPLC-grade reagent water at approximately
22°C and analyzed by high performance liquid chromatogr aphy. Based on the results, the test substance is
soluble at 8500 mg/L.REFERENCES

1. U.S. Environmental Protection Agency 1996. Water Solubility: Column Elution Method; Shake Flask Method. Product Property Test Guidelines. OPPTS 830.7840.

2. Organisation for Economic C ooperation and Development. 1995. Guideline for Testing of Chemicals, 105: Water Solubility.

3. TSCA Title 40 of the Federal Code of Regulations. 1994. Part 796, Section 1840: Water Solubility.Table 1

Typical HPLC Operational Parameters

INSTRUMENT: Agilent Model 1100 High Perfor mance Liquid Chromatograph (HPLC) equipped with an Ag ilent Series 1100 Variable Wavelength Detector

ANALYTICAL COLUMN: YMC-Pack ODS-AM (150 x 4.6 mm, 3- μ m particle size)

STOP TIME:

FLOW RATE:

SOLVENT A:

SOLVENT B: 10 minutes

1.000 mL/min

10% (0.1% H 3 PO 4 )

90% (CH 3 CN)

OVEN TEMPERATURE: 40°C

INJECTION VOLUME: 25.0 μ L

APPROXIMATE RETENTION TIMES: Component #1 – 4.4 minutes Component #2 – 5.5 minutes Component #3 – 7.8 minutes

WAVELENGTH: 200 nm- 14 -

Table 2

Solubility Results of the Test Substance in HPLC-Grade Reagent Water

Nominal Concentration (mg/L) Sample I.D. (523C-104-) Sampling Time (Day) Temp. at Sampling (°C) Measured Concentration (mg/L) 1,2 Percent of Nominal 1 Mean Measured Std. Dev. (SD) RSD. 1

8500 8500 8500

8500 8500 8500

8500 8500 8500 D1A D1B D1C

D2A D2B D2C

D3A D3B D3C 1 1 1

2 2 2

3 3 3 22.6

22.5

22.0 8109 8123 8062

8089 8297 8290

8107 8344 8332 95.4 95.6 94.9

95.2 97.6 97.5

95.4 98.2 98.0 Mean = 8098 SD = 31.6 RSD = 0.390%

Mean = 8225

SD = 118 RSD = 1.43%

Mean = 8261

SD = 134 RSD = 1.62%
Mean = Std. Dev. = RSD = 8195 117 1.43

1 Results were generated using Excel 2000 in the full precision mode. Manual calculations may differ slightly.

2 The limit of quantitation (LOQ) was 1665 mg/L, calculated as the product of the concentration of the lowest calibration standard (5.00 mg/L) and the dilution factor of the samples (333).- 15 -

Figure 1. Analytical method flowchart for the analysis of the test substance in HPLC-grade reagent
water.0 200 400 600 800 1000 1200

0. 00 10. 00 20. 00 30. 00 40. 00 50. 00 60. 00 Concentration (mg/L)Peak Area

Figure 2. A representative calibration curve for the test substance Slope = 20.63516; Y-Intercept = -9.28055; R 2 = 0.99991Figure 3. A representative chromatogram of a low-level (5.00 mg/L) test substance calibration standard.Figure 4. A representative chromatogram of a high-level (50.0 mg/L) test substance calibration standard.Figure 5. A representative chromatogram of a Day 1 water solubility sample (523C-104-D1B), nominal concentration 8500 mg/L.Appendix 1

Protocol, Protoc ol Amendments and Deviation- 24 -Appendix 2

Personnel Involved in the Study

The following key