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### 1 PROGRESS REPORT: MARCH 2012

BIODEGRADATION TESTING SUPPLEMENTAL ENVIRONMENTAL PROJECT

Submitted to

E.I du Pont de Nemours and Company

Wilmington, Delaware 19898

U.S.A.

United States Environmental Protection Agency

Washington, DC

U.S.A.



# Wildlife International, Ltd.

8598 Commerce Drive

Easton, Maryland 21601 USA

1 (410) 822

- 8600

March 28, 2012

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### 2 Table of Contents

Title Page

................................ ................................ ................................ ................................ ... 1

Table of Contents

................................ ................................ ................................ ....................... 2

Introduction

................................ ................................ ................................ ................................ 3

Progress Summary

................................ ................................ ................................ ..................... 3

Appendix A. Invoice

................................ ................................ ................................ ................. 4

Appendix B. LC/MS/MS Instrumental Conditions and Representative Chromatograms for

Calibration Standards of 6

- 2 FTOH, 8

- 2 FTOH and 10

- 2 FTOH

.................... 6

Appendix C. Calibration Standard Storage Stability Data (7 Days, Refrigerated)

................ 34

Appendix D. Ana lytical Standard, Stock Solution and Reagent Preparation Information

..... 40

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### 3 Introduction

This document presents the progress of the technical work for the Biodegradation Testing

Supplemental Environmental Project (“Biodegradation SEP”) performed from 27 Febru ary

2012 up to 23 March 2012. Studies conducted under protocol will be examined by the

Wildlife International Quality Assurance Unit for compliance with Good Laboratory

Practices, Standard Operating Procedures and the specified protocol. Raw data that ha s been

audited by the Wildlife International Quality Assurance will be included in the progress

report, as appropriate.

Progress Summary

The total costs associated with the project for the period covered in this Progress Report are

approximately $5,335 fo r labor and $18,900 for instrumentation. The invoice and an

allocation of costs with associated tasks are presented in Appendix A.

The technical work consisted of initiation of the SCAS Analytical Method Development Plan

(20 Jan 2012.doc) as approved by EPA on February 14, 2012. The objective of this work is

to determine if an LC/MS/MS method can be developed for determination of 6

- 2 FTOH, 8

- 2

FTOH, and 10

- 2 FTOH in MTBE extracted SCAS matrix.

LC/MS/MS instrumental conditions and calibration ranges hav e been established for

quantitation of all three telomer alcohols and their associated matrix verification, mass

labeled analogs (M+4 6

- 2 FTOH, M+4 8

- 2 FTOH, and 1 M+4 10

- 2 FTOH). Calibration

standards for 6

- 2 FTOH, 8

- 2 FTOH, and 10

- 2 FTOH, ranging in co ncentration from 5.00 to

250 ng/mL, were prepared in a solvent system consisting of 40:40:20 (v/v)

MTBE:ACN:H2O. Each calibration standard was prepared to contain 100 ng/mL in each of

the three mass

- labeled telomer alcohols. Appendix B provides currently employed

instrumental conditions, an example calibration curve and representative chromatograms for

each component in the calibration standard set. The standards have been shown to be stable

for a minimum of one week following initial preparation (Append ix C). Records of standard

and solution preparation associated with this work have been summarized in Appendix D.

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### 4 Appendix A

Invoice

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### 5 Allocation of Costs

Month, Year Tasks Total

Design and

Procurement

of SCAS Units

(Task A) Acquisition

of

Standa rds

(Task B) Adaptation of

Analytical

Method (Task C) Pilot Studies

Task (D) Refinement

of Methods

Task (E) Method

Val

(Task F) Storage

Stability

(Task G) Biod

Studies

(Task H) Instrument

Lease

Dec 2009 $1,803 $1,085 $42,422 $45,310

Jan 2010 $29,227 $29,227

Feb 2010 $1,300 $36,745 $38,045

Mar 2010 $14,350 $14,350 $18,900

Apr 2010 $570 $15,935 $16,505 $18,900

May 2010 $760 $71,860 $72,620 $18,900

June 2010 $57,870 $35,003 $92,873 $18,900

July 2 010 $42,430 $15,130 $57,560 $18,900

Aug 2010 $32,530 $113,718 $146,248 $18,900

Sep 2010 $1,350 $115,228 $116,578 $18,900

Oct 2010 $88,823 $3,613 $92,478 $18,900

Nov 2010 $42,690 $855 $570 $44,115 $18,900

Dec 2010 $ 18,385 $3,530 $3040 $24,955 $18,900

Jan 2011 $15,570 $15,570 $18,900

Feb 2011 $2,945 $2,945 $18,900

Mar 2011 $8,075 $8,075 $18,900

Apr 2011 $9,820 $9,820 $18,900

May 2011 $5,770 $5,770 $18,900

June 2011 $2,850 $190 $3,040 $18,900

July 2011 $2,965 $2,965 $18,900

Aug 2011 $950 $10,713 $11,663 $18,900

Sep 2011 $1,140 $1,140 $18,900

Oct 2011 $380 $950 $1,330 $18,900

Nov 2011 $1,235 $1,235 $18,900

Dec 2011 $1, 520 $1,520 $18,900

Jan 2012 $1,140 $1,140 $18,900

Feb 2012 $1,140 $1,140 $18,900

Mar 2102 $5,335 $5,335 $18,900

Labor Total $1,803 $3,715 $344,719 $430,318 $79,165 $3,800 $863,520

Materials $6,800 $10,500 $ 20,644 $1,724 $1,813 $41,482

Total Cost $8,603 $14,215 $365,363 $432,042 $80,978 $3,800 $905,002 $472,500

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### 6 Appendix B

LC/MS/MS Instrumental Conditions and Representative Chromatograms for Calibration

Standards of 6

- 2 FTOH, 8

- 2 FTOH and 10

- 2 FTOH

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Calibration Standard Storage Stability Data (7 Days, Refrigerated)

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Analytical Standard, Stock Solution and Reagent Preparation Information

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