References1 Klosterhaus et al. 2008. 10th Annual Workshop on Brominated Flame

Retardants. Victoria, British Columbia. June 3 and 4, 2008.

2 http://riodb01.ibase.aist.go.jp/sdbs/cgi-bin/direct_frame_top.cgi

3 Marklund et al. 2003. Chemosphere 53:1137.

4 Reemtsma et al. 2008. Trends in Analytical Chemistry 27: 727.

5 Stapleton et al. 2009. 11th Annual Workshop on Brominated Flame Retardants. Ottawa, Ontario. May 19 and 20, 2009.

6 Webster et al. 2009. Identifying Transfer Mechanisms and Sources of Decabromodiphenyl Ether (BDE 209) in Indoor Environments Using Environmental Forensic Microscopy. Environmental Science & Technology, ASAP Alerts.

7 Babich, M. A. 2006. CPSC Staff Preliminary Risk Assessment of Flame Retardant (FR) Chemicals in Upholstered Furniture Foam. US CPSC Bethesda, MD 20814.

8 US EPA. 2005. Furniture Flame Retardancy Partnership: Environmental Profiles of Chemical Flame Retardant Alternatives. 742-R-05-002A.

.

Firemaster 550

Time16.00 16.50 17.00 17.50 18.00 18.50 19.00 19.50 20.00 20.50

%

0

100

16.00 16.50 17.00 17.50 18.00 18.50 19.00 19.50 20.00 20.50

%

0

100

16.00 16.50 17.00 17.50 18.00 18.50 19.00 19.50 20.00 20.50

%

0

100

16.00 16.50 17.00 17.50 18.00 18.50 19.00 19.50 20.00 20.50

%

0

100 368.11771.21e8

15.90

16.75

16.33

410.16471.71e7

16.74

16.28

17.14

16.90

17.68

17.20

17.2618.2317.80 18.78 19.03

452.21164.89e6

18.04

17.75

17.62

16.96

19.1618.1418.62 18.82

19.32 19.86

494.25864.67e5

19.00

18.65

19.50

19.16 20.21

19.55

19.62 20.26

Mono-ipPP

Tetra-ipPP

Tri-ipPP

Peak Calc % of Retention Time Conc Total

Assuming AssumingEqual Response Equal Response

pg/µg (ppm)Mono-ipPP

15.90 19938616.33 4148216.75 82157

Total Mono-ipPP 323025 31.9

Di-ipPP16.28 45616.74 2766916.90 648417.15 2628517.20 819817.26 640517.68 1774017.80 142418.06 61218.23 295418.78 334619.03 166

Total Di-ipPP 101741 10.1

Tri-ipPP

16.96 29217.62 149017.75 330918.04 897418.14 151218.27 43618.59 110718.62 159818.75 55518.82 55419.16 244619.23 40919.32 66719.87 441

Total Tri-ipPP 23788 2.4

Tetra-ipPP

18.65 41818.77 5418.81 3019.00 86719.16 47919.23 8619.26 5919.29 8319.50 85919.55 20619.62 21819.82 17320.21 33720.26 74

Total Tetra-ipPP 3942 0.4Total mono+di+tri+tetra 452496

0.45

Other ComponentsTPP 172000 17.0T2iPPP* 1490 0.1TBB 300000 29.6TBPH 87500 8.6Total Other Components 559500

Grand Total 1011996 101

* Already counted

Di-ipPP

min

557,423

12.0 20.0 30.0 40.0 0

50000

100000

150000

200000

250000

300000

350000

400000

450000

500000

550000

21.3

03

22.8

17

24.2

1724

.782

28.1

12

OP

O

OOOP

O

OO

OP

O

OO

OP

O

OO

tri o-tolyl phosphate (TOTP)

triphenyl phosphate(TPP)

tris(3,5-dimethylphenyl) phosphate (TDMPP)

tris(2-isopropylphenyl) phosphate (TIPP)

tris(4-tert-butylphenyl) phosphate (TTBPP)

OP

O

OO

GC-MS (EI) of �ve aryl phosphates at 10 ug/ml.

RRF FW Corrected

TPP 1.0 1.0

TOTP 0.9 1.0

TIPP 0.9 1.2

TDMPP 0.6 0.7

TTBPP 0.1 0.2

Relative Response Factors:

intensity9,461,267

19.0 20.0 21.0 22.0 23.0 24.0 25.0 26.0 27.0 28.0 0

5000000

TIC*1.00

19.7

0 TP

P

20.7

9 m

/z 3

68: 2

-iPPP

21.2

9 m

/z 3

68: 4

-iPPP

21.7

4 m

/z 3

68: 3

-iPPP

+m/z

410

21.9

5 m

/z 4

10

22.1

1 m

/z 4

5222

.21

m/z

410

22.3

6 m

/z 4

10

22.6

1 m

/z 4

52 tr

is-2

iPPP

22.7

1 EH

TBB

22.7

8 m

/z 4

52

23.0

3 m

/z 4

5223

.16

m/z

452

23.2

5 m

/z 4

52

23.4

9 m

/z 4

5223

.57

m/z

410

23.6

7 m

/z 4

5223

.79

m/z

494

23.9

2 m

/z 4

5223

.97

m/z

494

24.0

6 m

/z 4

94

24.2

1 m

/z 4

94

24.3

7 m

/z 4

9424

.48

Hal

ogen

ated

unk

now

n

24.6

3 m

/z 4

94

24.8

9 H

alog

enat

ed u

nkno

wn

27.2

3 BE

HTB

P

O PO

OO

O PO

OO

O PO

OO

O PO

OO

O PO

OO

m/z 368 - mono-isopropylated Triphenyl Phosphatem/z 410 - di-isopropylated Triphenyl Phosphatem/z 452 - tri-isopropylated Triphenyl Phosphatem/z 494 - tetra-isopropylated Triphenyl Phosphate

BrBr

BrBr

O

O

O

O

BrBr

BrBr

O

O

TIC*1.00

SpinWorks 2.5: FM550T2 in CD2Cl2 - JMOD - Quaternary and CH2 up, CH3 and CH down

PPM38 36 34 32 30 28 26 24 22 20 18 16 14 12

33.

9628

33.

5497

26.

7539

23.

7652

23.

5397

22.

5567

22.

5082

OP

O

O

O

a

a

b

b

c

c

d

d

d

f

f

f

e

e

e

OP

O

O

O

2-isopropylphenyl diphenyl phosphate 3-isopropylphenyl diphenyl phosphate 4-isopropylphenyl diphenyl phosphate

OP

O

O

O

TPP at WWTP #1: bar represents mean and standard deviation of three replicates

TBB, TBPH, BDE 209, total PBDE at WWTP #1, TPP at WWTP #2: bars represent mean of two replicates

TBB, TBPH, BDE 209, total PBDE at WWTP #2: bars represent concentration in one sample

Components of Firemaster 550®

ng

/g d

ry w

eig

ht

JMOD Spectra of Firemaster 550® TLC Band 2. Aliphatic Region Identified Components in Firemaster 550®

Firemaster 550® Quantification Results

Relative Response Factors of Some Aryl Phosphates

Concentrations of Flame Retardant Chemicals in Biosolids Collected from San Francisco Bay Area Wastewater Treatment Plants

FIGURE

4

FIGURE

5

FIGURE

3FIGURE

2FIGURE

1

Susan Klosterhaus 1, Alex Konstantinov 2, Elizabeth Davis3, Jeffery Klein2, and Heather Stapleton 3

1 San Francisco Estuary Institute, Oakland, CA, USA.2 Wellington Laboratories, Guelph, Ontario, Canada.3 Duke University, Nicholas School of the Environment & Earth Sciences, Durham, NC, USA.

Characterization of Organophosphorus Chemicals in a PentaBDE Replacement Mixture and their Detection in Biosolids

AcknowledgementsWe thank Chemtura for providing the sample of Firemaster 550®

Contact InformationSusan Klosterhaus: susan@sfei.org

Alex Konstantinov: alex@well-labs.com

Elizabeth F. Davis: elizabeth.f.davis@duke.edu

Jeffery Klein: jeff@well-labs.com

Heather Stapleton: heather.stapleton@duke.edu

Firemaster 550® Chemical Analyses Triphenyl phosphate (TPP) was purchased from Aldrich. Tri o-tolyl phosphate was purchased from Fisher Scientific. Tris(2-isopropylphenyl) phosphate (TIPP), tris(3,5-dimethylphenyl) phosphate, and tris(4-tert-butylphenyl) phosphate were pur-chased from ChemBridge Corporation. 2-Ethyl-1-hexyl 2,3,4,5-tetrabromobenzoate (TBB) and bis(2-ethyl-1-hexyl) tetrabromophthalate (TBPH) were from Wellington Laboratories. A sample of Firemaster 550® was provided by Chemtura.

Low resolution electron impact (EI) analyses were performed on a Shimadzu GC/MS-QP2010 using a J&W 30m DB-5 column (0.25 mm ID,

0.25 µm film). All injections were done in split-less mode. All experiments were performed using the following GC conditions:

• Helium carrier gas flow at 1.0 ml/minute.

• Injector temperature at 250°C.

• GC oven temperature program: initial oven temperature was 100°C for 5 minutes; 10°C/minute to 325°C ; hold for 20 minutes.

• Spectra (50 to 1000 u) were obtained in positive ion, electron impact mode (EI+).

High resolution analyses were performed on Agilent 6890N HRGC / Waters Autospec Ultima HRMS equipped with a J&W 60 m DB-5 (0.25 mm ID, 0.25 µm film) column.

The following conditions were used:

• Helium carrier gas with constant flow at 1.0 ml/minute.

• Injector, source and transfer line at 280°C.

• GC oven temperature program: initial oven temperature was 100°C for 2 minutes; 25°C/minute to 250°C; 10°C/minute to 325°C; hold for 15 minutes.

• Data were acquired using SIM monitoring at a resolution of 10,000 in EI+ mode.

1H-NMR analyses were performed on a 600 MHz Bruker instrument using deuterated methylene chloride (CDN isotopes) as the solvent.

Characterization of Organophosphorus Compounds in Firemaster 550®

Indentification: At least 24 alkylated phenyl phosphates were present in the FM-550 technical mixture according to gas chromatography-mass spectrometry (GC-MS). To make identification feasible, a large sample of FM-550 was subjected to preparative TLC separation. Separation yielded five TLC bands. The band with the lowest Rf value was triphenyl phosphate (TPP; 14% by weight), followed by three bands containing mono-, di-, and tri- isopropylphenyl phosphates (iPPs), respectively. The fifth TLC band (46% by weight) contained the brominated components of the mixture: 2-ethyl-1-hexyl 2,3,4,5-tetrabromobenzoate (TBB) and bis(2-ethyl-1-hexyl) tetrabro-mophthalate (TBPH).

In addition to the analysis of 1H NMR data, analysis of TLC bands two, three and four by 13C NMR (JMOD experiment) indicated an absence of CH2 signals and isolated CH3 groups, which is consistent with isopropyl-only alkyl groups. Examination of 13C chemical shifts for TLC band two allowed for identification of ortho-, meta- and para- mono isopropyl isomers in FM-550 (FIGURE 1). The assignments were made by com-parison of the chemical shift values for the methyl and isopropyl carbons to those of the corresponding isopropyl phenols in the NMR Library (2). The assignments were additionally confirmed by 1H NMR data comparisons. The ortho isomer was the major isomer, followed by meta and para isomers.

For the remaining bands, identification by NMR analysis was not possible as high structural similarity resulted in signal overlaps. Additionally, tris (2-isopropylphenyl) phosphate (T2iPP) was identified as a minor component in FM-550 (FIGURE 2).

Quantification: Relative response factors (RRFs) for various alkylated phenyl phos-phates on GC-MS were also measured. The RRFs for TPP and T2iPP were found to be very similar and, in the absence of other isopropylated phenyl phosphate standards, the isopropyl aryl phosphates in FM 550 were quantified using commercially available T2iPP as a standard. (FIGURE 3).

Using a four point calibration (BDE-126 as an injection standard; RSD lower than 10%) and assuming an equal response for all isopropylated phenyl phosphates, the concentrations of the components of FM 550 were: TPP – 18%, Mono-iPP – 32%, Di-iPP – 10%, Tri-iPP – 2.4%, Tetra-iPP – 0.4%, TBB – 29.6%, and TBPH – 8.6%. There were also several co-elutions obvious from High Resolution GC-MS data (FIGURE 4).

IntroductionWorldwide restrictions on the use of PentaBDE have led to the use of new flame retardant mixtures to meet flammability standards. Assessments to determine if the chemicals in these mixtures are accumulating in indoor and outdoor environments have thus far not been possible because their structural identities are not readily available. In our study, the phosphate components of Firemaster 550® (FM-550), a commercial mixture used in high volumes to meet the California furniture flammability standard, were characterized and quantified in biosolids collected from two municipal wastewater treatment plants (WWTPs) that discharge effluent to San Francisco Bay. The analysis of biosolids was conducted to determine if these compounds are migrating out of consumer products. This study augments previous work which identified the brominated chemicals in FM-550 (1).

Organophosphorus Compounds in BiosolidsTPP was detected in the biosolids in concentrations ranging from 530-2630 ng/g dry weight, with mean concentrations of 2048 ± 544 and 945 for each of the WWTPs (FIGURE 5). AT WWTP #1, the ratio of TPP to the other FM 550 components (TBB and TBPH) was fairly similar to the ratio found in the commercial mixture (FIGURE 4). However, for WWTP #2, the mean TPP concentration was higher than TBB. TPP is a high volume chemical which has been used for decades as a plasticizer in several appli-cations (3); thus FM-550 is not the only potential source of TPP to the biosolids. At both WWTPs, TPP was comparable in concentration to BDE 209. Detection in biosolids suggests that TPP, a moderately hydrophobic compound (log Kow 4.59) (4), has the po-tential to be released to the environment via wastewater effluent and accumulate in aquatic environments.

Implications for Organophosphorus Compounds in the EnvironmentIn addition to the detection of TPP in biosolids in this study, TPP was also recently found in house dust at concentrations similar or higher than PBDEs (5). It has also been detected in indoor air, wastewater effluents, and surface waters (3). This information suggests that TPP can migrate out of consumer products, similar to BFRs (6).

The US Consumer Product Safety Commission has concluded that both TPP and iPPs are ‘possibly toxic’ to humans based on limited evidence of neurotoxicity in animal studies (7). US EPA considers TPP a high hazard concern for ecotoxicity (8). Potential exposure to humans and wildlife warrants further assessment of TPP environmental fate and toxicity due to the potential increased use of TPP in flame retardant mixtures as a replacement for PentaBDE.

Biosolids Analyses • Collected in February 2008 from two

WWTPs that discharge effluent to San Fran-cisco Bay

• Extracted using pressurized fluid extraction (3X with 100% dichloromethane)

TBB, TBPH, and PBDE Extract Clean-up and Analysis

• Extracts purified using 8.0 g of 2.5% deacti-vated Florisil; rinsed with 50 mL of 50:50 hexane:dichloromethane.

• Analyzed using GC/ECNI-MS; 0.25 mm (I.D.) x 15 m fused silica capillary column coated with 5% phenyl methylpolysiloxane (0.25 µm film thickness).

• PTV injection; inlet temperature 50 °C for 0.3 minutes and then a 700 °C/min ramp to 275 °C.

• GC oven temperature program: hold at 40 °C for 1 min, ramp of 18 °C /min to 250 °C,

ramp of 1.5°C/min to 260 °C, ramp of 25 °C/min to 300 °C, 20 min. hold.

• Transfer line and ion source temperature: 300 °C and 200°C respectively.

• Molecular fragments quantified: m/z 484.6 and 486.6 (BDE 209); m/z 494.6 and 496.6 (13C BDE-209); m/z 357 (Quantitative) and 471 (Qualitative) (TBB); m/z 463 (Quantitative) and 515 (Qualitative) (TBPH).

• Quantification standards: F-BDE-160 and 13C-BDE-209.

TPP Extract Clean-up and Analysis

• Extracts purified using 4.0 g of 6.0% deacti-vated alumina; rinsed with 50 mL of 50:50 hexane:dichloromethane.

• Extracts blown to dryness under a N2 stream, reconstituted in dichloromethane.

• Purified by using a GPC column (EnvirogelTM GPC Cleanup 90x300 mm column, dichloromethane mobile phase, flow rate of

5 mL/min). The first 60 mL of eluate were discarded, and the following 40 mL were collected.

• Extract was solvent-switched to hexane.

• Quantification standard: 13C-CDE 141.

• Analyzed using GC/MS (EI); 0.25 mm (I.D.) x 15 m fused silica capillary column coated with 5% phenyl methylpolysiloxane (0.25 µm film thickness).

• PTV injection; inlet temperature 80 °C for 0.3 minutes and then a 600 °C/min ramp to 300 °C.

• GC oven temperature program: hold at 80 °C for 2 min, ramp of 20 °C /min to 250 °C, ramp of 1.5°C/min to 260 °C, ramp of 25 °C/min to 300 °C, 12 min. hold.

• Transfer line and ion source temperature: 300 °C and 200°C respectively.

• TPP was quantified using ion fragments m/z 326 (quantitative) and 325 (qualitative).

Materials and Methods

R M P P I L OT A N D S P E C I A L S T U D I E SREGIONAL MONITORING PROGRAM FOR WATER QUALITY IN THE SAN FRANCISCO ESTUARY