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2011_July_Progress Report 1

PROGRESS REPORT: JULY 2011 

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 

July 29, 2011 

Page 1 of 6 
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2011_March _Progress Report 2 

Table of Contents 

Title Page ...................................................................................................................................1 

Table of Contents.......................................................................................................................2 

Introduction................................................................................................................................3 

Progress Summary .....................................................................................................................3 

Appendix A. Invoice.................................................................................................................4 

Appendix B. Task E: Modifications and Refinements to Test Methods .................................6 
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2011_March _Progress Report 3 

Introduction 

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

Supplemental Environmental Project (“Biodegradation SEP”) performed from 25 June 2011 

up to 23 July 2011. 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 has 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 $2,965 for labor and $18,900 for instrumentation. The invoice and an 

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

Work conducted over the period covered by this progress report consisted of the review and 

discussion of data from test substance characterization and discussion of Task E items with 

stakeholders. Work included revision of the Scope of Work (SOW) presented in the June 

2011 Progress Report. The SOW was composed to investigate approaches to eliminate or 

avoid carryover and enhancement effects observed and documented for GC/MS 

determinations of 6-2 FTOH, 8-2 FTOH and 10-2 FTOH in MTBE extracts during the initial 

pilot study with Group A1 polymer. The initial SOW document presented an outline for 

developing an LC/MS/MS method for these alcohols as a potential solution to the problem. 

Another approach presented in the interim is use of increased centrifugation force and time to 

minimize or eliminate co-extracted polymer in the MTBE extract as a prelude to the original 

GC/MS determination method. The SOW was therefore revised to include this experimental 

option (Appendix B). 
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2011_March _Progress Report 4 

Appendix A 

Invoice 
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2011_July_Progress Report 

Allocation of Costs 

Month, Year Tasks 

 Design and 

Procurement of 

SCAS Units 

(Task A) 

Acquisition 

of Standards 

(Task B) 

Adaptation of 

Analytical 

Method (Task C) 

Pilot Studies 

Task (D) 

Refinement 

of Methods 

Task (E) 

Method 

Validations 

(Task F) 

Storage 

Stability 

(Task G) 

Biod

dat

Stu

(Tas

Dec 2009 $1,803 $1,085 $42,422 Jan 2010 $29,227 Feb 2010 $1,300 $36,745 Mar 2010 $14,350 Apr 2010 $570 $15,935 May 2010 $760 $71,860 June 2010 $57,870 $35,003 July 2010 $42,430 $15,130 August 2010 $32,530 $113,718 September 2010 $1,350 $115,228 October 2010 $88,823 $3,613 November 2010 $42,690 $855 $570 December 2010 $18,385 $3,530 $3040 January 2011 $15,570 February 2011 $2,945 March 2011 $8,075 April 2011 $9,820 May 2011 $5,770 June 2011 $2,850 $190 July 2011 $2,965 Labor Total $1,803 $3,715 $344,719 $428,988 $55,993 $3,800 Materials $6,800 $10,500 $20,644 $1,724 Total Cost $8,603 $14,215 $365,363 $430,712 $55,993 $3,800 9
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2011_July_Progress Report 6

Appendix B. Task E: Modifications and Refinements to Test Methods 

From the document Task E: Refinement of Methods (2010_November_02_Task 

E_detail.doc), Item 3 is the only remaining analytical method issue identified by Wildlife 

International, Ltd. following the initial pilot study not yet resolved following consultation 

with DuPont/EPA. The original statement is reproduced below (with minor 

clarifications/corrections indicated in parenthesis): 

3) After injection of sludge extracts from polymer treatment SCAS chambers, an enhanced 

(GC/MS) signal for 6-2 FTOH, 8-2 FTOH, and 10-2 FTOH (was) observed and the 

enhanced signal persisted for subsequent injections of standards. 

The consensus of the stakeholders is that the MTBE activated sludge extracts contain 

polymer co-extractives, which are degrading under the thermal conditions of the GC/MS 

inlet to form the alcohols. One approach is to migrate from GC/MS to LC/MS/MS for 

determination of 6-2 FTOH, 8-2 FTOH, and 10-2 FTOH in MTBE extracts. 

A. Fortify replicate negative control sludge matrices with 

1. 6-2 FTOH, 8-2 FTOH and 10-2 FTOH 

2. Fluorotelomer Product Group A1 polymer 

3. A1 polymer and 6-2 FTOH, 8-2 FTOH and 10-2 FTOH 

B. Process following the sample preparation and LC/MS/MS instrumental conditions 

presented in Biodegradation SEP, Attachment D, Analytical Methods: Szostek, B.; 

Prickett, K.B.; Buck, R.C., Determination of fluorotelomer alcohols by liquid 

chromatraphy/tandem mass spectrometry in water. Rapid Communications in Mass 

Spectrometry 2006, 20, (19), 2837-2844. 

 

1. Perform infusion experiments using LC/MS or LC/MS/MS to evaluate the 

feasibility of performing adduct free analyses in our facilities. 

2. If it is not viable to monitor deprotonated parent ions using LC/MS/MS due to 

the prevalence of formic/acetic acid residuals in our systems, develop 

alternative strategies using appropriate adducts of the three target alcohol 

analytes. 

3. Use LC/MS or LC/MS/MS depending on the requisite sensitivity and 

selectivity requirements to produce an appropriate and robust method. 

4. Investigate a combination of deprotonated parent ion and adduct monitoring in 

order to minmize instrumental setup time for future pilot and definitive SCAS 

trials. 

Estimated budget: $30,000 

Estimated time frame: 4 weeks

Another approach is to centrifuge the MTBE: reagent water solvent partion created during 

the back-partition MTBE extract clean-up step in the proposed activated sludge mixed liquor 

method at a higher relative centrifugal force and for a longer duration than the settings used 
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2011_March _Progress Report 7 

in the initial pilot study for sludge solids and supernatant SCAS matrices. The objective is to 

physically remove polymer co-extractives from the final cleaned MTBE extract prior to the 

GC/MS determination. 

A. Fortify replicate reagent water samples with variable amounts of Fluorotelomer 

Product Group A1 polymer corresponding to the range of polymer mass 

calculated to have been theoretically present in 10-mL mixed-liquor subsamples 

during the latter stages of the initial pilot study. Prepare replicate fortifications 

and perform sequential “A” and “B” MTBE extractions for each of the 

combinations of polymer mass and centrifugation force (during back-partition 

step) to be investigated per the most current activated sludge mixed liquor method 

outline. 

B. Transfer replicate aliquots from each MTBE extract into separate plastic 

centrifuge tubes containing reagent water for a final solvent ration of 1 part 

MTBE: 2 parts H2O and centrifuge at selected centrifugation force settings (3800 

xg and greater) for 30 minutes. Note: pre-weigh the empty plastic centrifuge 

tubes and use a common centrifugation force for any given pairing of “A” and 

“B” MTBE extracts. 

C. For each sample, make physical observations for evidence any polymer 

precipitate and take aliquouts of the supernatant for GC/MS determination of 6-2 

FTOH, 8-2 FTOH and/or 10-2 FTOH to measure any potential relative 

differences in alcohol response. Prepare post-spike samples of the desired 

telomer alcohol(s) as needed. 

D. Weigh the containers after processing for potential quantitation of any precipitate 

that may or may not be apparant in the plastic vessels. 

Estimated budget: $8,000 

Estimated time frame: 1 week