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QUANTITATIVE ANALYSIS AND HEALTH RISK

ASSESSMENT OF NOVEL BROMINATED FLAME

RETARDANTS IN HOUSE DUST

Taya Huang

MSc Thesis

Master's Degree Programme in Environmental Health Risk Assessment

University of Eastern Finland, Faculty of Science and Forestry

Department of Environmental and Biological Sciences

28 April, 2017

UNIVERSITY OF EASTERN FINLAND, Faculty of Science and Forestry

Department of Environmental Science

Master's Degree Programme in Environmental Health Risk Assessment

Taya Huang: Quantitative Analysis and Health Risk Assessment of Novel Brominated Flame

Retardants in House Dust

MSc thesis 66 pages, 8 appendixes ( 74 pages)

Supervisors: Panu Rantakokko, PhD; Matti Viluksela, PhD

28 April, 2017

________________________________________________________________________

Keywords: flame retardants, brominated flame retardants, method development,

environmental health risk assessment, indoor dust

ABSTRACT

This thesis discusses the method development and validation for analysis of flame retardants

in house dust that was performed between May-August 2015 based on the method developed

by Van den Eede et al. 2012. The principle aim of this method development is to develop and

validate an effective analysis method for indoor dust containing selected novel and emerging

brominated flame retardants (BFR), organophosphorous flame retardants (OPFR), as well as

polybrominated diphenyl ether (PBDE). In the second part of this thesis, a human health risk

assessment for one of the above selected BFRs, namely Bis(2-ethyl-1hexyl)

tetrabromophthalate (BEH-TEBP) is performed based on currently available scientific

literature and estimated exposure from indoor dust ingestion. For the validation, Limit of

Quantification (LOQ) & Measurement Uncertainties were acceptable for all BFRs. LOQ of

BFRs ranged between 0.5 – 5.0 ng/g. For OPFR, LOQ ranged between 6.9-613 ng/g.

Precision for OPFR was good, but accuracy was poor compared to other labs’ analysis result

for the certified material for indoor dust SRM 2585. Therefore, method for OPFR analysis

requires further testing that was not conducted within the scope of this thesis. For the purpose

of estimating exposure, unpublished results from the National Institute for Health and Welfare

(THL) were used. The detected amount of BEH-TEBP from children’s room in Kuopio,

Finland (n=40) had a median of 106.3 ng/g, with a range of 22.8 ng/g – 887.2 ng/g (5th to 95th

percentile). A Risk Characterisation Ratio (RCR) has been calculated based on the derived

no-effect level (DNEL) of 0.37 mg/kg bw/day for long-term oral exposure in the general

population, taken from the registration dossier of BEH-TEBP. All RCR derived for given

exposure scenarios are less than 1, meaning that the risk is adequately controlled. However, it

must be kept in mind that combined exposure has not been taken into consideration.

ACKNOWLEDGEMENTS

The laboratory work for this thesis was performed between May-August 2015. The method

development and validation for analysis of flame retardants in house dust described in this

thesis has been done according to the instructions and guidance from Dr. Panu Rantakokko of

the National Institute for Health and Welfare (THL), Kuopio. The official validation report for

the first validation has been written by Dr. Rantakokko for THL. The method development

was still on-going as of August 2016. This thesis includes only the part of the work I have

been involved in. This laboratory work forms the first part of my thesis. The second part of

my thesis focuses on the environmental health risk assessment of selected brominated flame

retardants.

ABBREVIATIONS AND DEFINITIONS

Terms

BFR Brominated Flame Retardant

DNEL Derived No-Effect Level

FR Flame Retardant

LOAEL Lowest Observed Adverse Effect Level

LOQ Limit of Quantification

NOAEL No Observed Adverse Effect Level

NOEL No Observed Effect Level

OPFR Organophosphorous Flame Retardant

RCR Risk Characterisation Ratio

REACH European Commission Regulation EC

1907/2006 concerning the Registration,

Evaluation, Authorisation and Restriction of

Chemicals

THL National Institute for Health and Welfare

USEPA Unites States Environmental Protection

Agency

Flame Retardants

ab-DBE-DBCH alpha/beta-Tetrabromoethylcyclohexane

BEH-TEBP Bis(2-ethyl-1hexyl)tetrabromophthalate

BTBPE 1,2-Bis(2,4,6-tribromophenoxy)ethane

DEHP di-2-ethyl hexyl phthalate

EHDPP 2-Ethylhexyl diphenyl phosphate

EH-TBB 2-Ethylhexyl-2,3,4,5-tetrabromobenzoate

PBDE Polybrominated diphenyl ether

PBT Pentabromotoluene

TBOEP Tris(2-butoxyethyl)phosphate

TBP-DBPE 2,3-Dibromopropyl 2,4,6-tribromophenyl ether

TCEP Tris(2-chloroethyl) phosphate

TCIPP Tris(1-chloropropan-2-yl) phosphate

TDBPP Tris(2,3-dibromopropyl) phosphate

TDCIPP Tris(1,3-dichloroisopropyl) phosphate

TEHP Tris(2-ethylhexyl)phosphate

TIBP Tri(isobutyl)phosphate

TMPP Tris (methylphenyl) phosphate (isomer mixture)

TNBP Tri-n-butylphosphate

TPHP Triphenyl phosphate

Chemicals

DCM Dichloromethane

HEX Hexane

Contents

1. INTRODUCTION 6

2 ANALYTICAL METHOD DEVELOPMENT FOR BFRs IN HOUSE DUST 12

2.1 MATERIALS AND METHODS 12

2.1.1 CHEMICALS AND INSTRUMENTS 12

2.1.2 GC-HRMS CONDITIONS 13

2.1.3 FRACTIONATION AND CLEAN-UP 14

2.1.4 EXTRACTION 20

2.1.5 VALIDATION 20

2.2 RESULTS AND DISCUSSION 26

2.2.1 FRACTIONATION AND CLEAN-UP OPTIMIZATION 26

2.2.2 EXTRACTION 27

2.2.3 SUMMARY OF SAMPLE TREATMENT IN FLOW CHART 29

2.2.4 RESULTS OF VALIDATION 29

2.2.5 CONCLUSIONS OF METHOD DEVELOPMENT AS OF 2016 34

2.2.7 DISCUSSION 35

3.1 BTBPE 39

3.1.1 Hazard Identification for BTBPE 39

3.1.2 Toxicity and Dose-Response Information on BTBPE 40

3.2 EH-TBB 40

3.2.1 Hazard Identification for EH-TBB 41

3.2.2 Toxicity and Dose-Response Information for EH-TBB 42

3.3 BEH-TEBP 43

3.3.1 Hazard Identification for BEH-TEBP 44

3.3.2 Toxicity and Dose-Response Information for BEH-TEBP 45

4 EXPOSURE ASSESSMENT 49

4.1 Available Relevant Exposure Information 49

4.2 Brominated Flame Retardants Measured in Children’s Room in Kuopio, Finland 50

5 RISK CHARACTERISATION FOR BEH-TEBP 57

6. DISCUSSION 57

7. SUMMARY AND CONCLUSIONS 61

8. REFERENCES 63

APPENDICES 67

APPENDIX 1: Reference values for BFR and OPFR analysis in house dust 67

APPENDIX 2: Criteria Used by USEPA to Assign Hazard Designation 69

6

1. INTRODUCTION

Flame retardants (FRs) are substances that can be added to polymer-based materials to

increase the materials’ resistance to ignition, and to slow down combustion. The purpose of

FRs is to reduce the risk of fire and delay the spread of fire. FRs are widely used in

commercial and consumer products due to existing fire safety regulations. Flammability tests

are required in building materials, industrial electronics and transportation vehicles

worldwide. For example, the EU Construction Products Directive stipulates flammability

standards for building materials. In the EU, the UK has set flammability standards for home

furniture. In Finland, there are flammability standards for upholstered furniture and mattresses

(EFRA 2015).

An example of required flammability standard in Finland is the EN 1021-1 & 2:2014, Decree

479/1996 for furniture upholstery. In a flammability standard test, the test fabric is exposed to

different ignition sources, specifically, a burning cigarette and butane flame, to examine its

burning behavior. The test material passes if no ignition occurs or there is only a limited area

of charring (SP 2016). As a result of flammability standards, flame retardants are ubiquitous

in the indoor environment, having been added to a wide range of industrial and commercial

products. FRs have been used in building insulation, furniture upholstery, carpet padding,

plastic casings for some electronics, and some baby products.

FRs can be emitted from these products into indoor air. The chemicals then bind to and settle

with indoor dust. FRs can accumulate in indoor dust, and it has been found that house dust

contributes to a high proportion of exposure in the indoor environment. Therefore, levels of

FRs present in house dust can be an indication of human exposure to FRs (Ni et al. 2013).

Moreover, concentration of FRs in gas phase can be estimated from measured concentrations

in house dust. For more volatile FRs, such as Pentabromotoluene (PBT), exposure through

gas phase may be significant as well (Little et al 2012).

Halogenated flame retardants and organophosphorous flame retardants make up approx. 30%

of FR used in the EU (European Union, 2011). Many of these FRs are known to be toxic,

persistent, bio-accumulative and can be transported through long distances. One example is

Polybrominated diphenyl ethers (PBDE). TetraBDE, pentaBDE, hexaBDE and heptaBDE

7

have been listed in the Annex A of the Stockholm Convention on Persistent Organic

Pollutants, whereby the production and usage of these mixtures have to be eliminated by the

parties to the Convention, for whom the amendment applies (United Nations 2015). In the

European Union, sale of commercial pentaBDE and octaBDE mixtures, in concentrations

higher than 0.1% by mass, has been banned. Under EU Directive 2002/95/EC, all new

electronic equipment must be free of PBDEs since July 2006. DecaBDE has been regulated

under Annex XVII of the European Commission Regulation EC 1907/2006 concerning the

Registration, Evaluation, Authorisation and Restriction of Chemicals (REACH), which

prohibits the use pf DecaBDE as a substance and restricts the content of DecaBDE in articles

to 0.1% (SGS 2017). All PBDEs are being regulated under the EU Restriction of Hazardous

Substances (RoHS) directive (European Union, 2011).

New BFR and OPFR mixtures have been developed to replace banned PBDEs. For example,

FM-550 and FMBZ-54 contain brominated and organophosphorous flame retardants such as

2-ethylhexyl-2,3,3,5-tetrabromobenzoate(EH-TBB) and bis-(2-ethylhexyl)-tetrabromophtalate

(BEH-TEBP). However, the toxicity of these novel and emerging FRs to human health, and

possible tendency for bio-accumulation have become a concern (Norrgran et al. 2015).

Determination of brominated and organophosphorous flame retardants in indoor dust

discussed in this thesis is based on the method developed by Van den Eede et al. 2012. The

principle aim of this method development is to develop and validate an effective analysis

method for indoor dust containing selected novel and emerging BFR, OPFR, as well as

PBDEs. Table 1 shows the BFR and OPFR that has been selected for the method

development.

8

Table 1. List of flame retardants included in method development.

CAS Number Abbreviation Full Chemical Name Molecular Weight Molecular Formula Chemical Structure

1 115-96-8 TCEP Tris(2-chloroethyl) phosphate 285.49 C6H12Cl3O4P

2 13674-84-5 TCIPP Tris(1-chloropropan-2-yl) phosphate 327.56 C9H18Cl3O4P

3 13674-87-8 TDCIPP Tris(1,3-dichloroisopropyl) phosphate 430.9 C9H15Cl6O4P

4 126-72-7 TDBPP Tris(2,3-dibromopropyl) phosphate 697.61 C9H15Br6O4P

5 126-71-6 TIBP Tri(isobutyl)phosphate 266.32 C12H27O4P

6 126-73-8 TNBP Tri-n-butylphosphate 266.32 C12H27O4P

9

7 78-51-3 TBOEP Tris(2-butoxyethyl)phosphate 398.48 C18H39O7P

8 1241-94-7 EHDPP 2-Ethylhexyl diphenyl phosphate 362.4 C20H27O4P

9 78-42-2 TEHP Tris(2-ethylhexyl)phosphate 434.64 C24H51O4P

10 115-86-6 TPHP Triphenyl phosphate 326.29 C18H15O4P

11 1330-78-5 TMPP Tris (methylphenyl) phosphate (isomer mixture) 368.37 C21H21O4P

12 3322-93-8 ab-DBE-

DBCH

alpha/beta-Tetrabromoethylcyclohexane 427.8 C8H12Br4

13 87-83-2 PBT Pentabromotoluene 486.62 C7H3Br5

10

14 35109-60-5 TBP-DBPE 2,3-Dibromopropyl 2,4,6-tribromophenyl ether 530.67 C9H7Br5O

15 183658-27-7 EH-TBB 2-Ethylhexyl-2,3,4,5-tetrabromobenzoate 549.92 C15H18Br4O2

16 37853-59-1 BTBPE 1,2-Bis(2,4,6-tribromophenoxy)ethane 687.64 C14H8Br6O2

17 26040-51-7 BEH-TEBP Bis(2-ethyl-1hexyl)tetrabromophthalate 706.14 C24H34Br4O4

11

In the second part of this thesis, a human health risk assessment for one of the above selected

BFRs, namely BEH-TEBP is performed based on currently available scientific literature and

estimated exposure from indoor dust ingestion. Risk assessment is based on the following

paradigm: hazard identification, dose-response assessment, exposure assessment and risk

characterization.

The OECD eChem portal, HSDB database and the ECHA CHEM database were consulted for

available relevant information on the above-listed BFRs, namely, ab-DBE-DBCH, PBT, TBP-

DBPE, EH-TBB, BTBPE and BEH-TEBP.

There was insufficient toxicological information available for ab-DBE-DBCH, PBT, TBP-

DBPE to perform a hazard identification. Relevant information available for the substances

BTBPE, EH-TBB and BEH-TEBP will be summarized in Section 3. The method described in

this thesis was used for the determination of BFRs from house dust samples collected around

Kuopio. An exposure estimation has been performed based on the amount of BFR detected

from a recent unpublished study by THL, where 40 house dust samples were analysed for

BFRs. Based on this exposure estimation, Risk Characterisation Ratio based on the Guidance

on Information Requirements and Chemical Safety Assessment (ECHA 2016a) has been

calculated for BEH-TEBP according to the DNEL for oral exposure.

The substance Bis(2-ethyl-1hexyl)tetrabromophthalate (BEH-TEBP) with CAS Number

26040-51-7 was therefore selected for risk characterization based on the availability of a

chemical registration dossier (ECHA 2016b) for BEH-TEBP through the European Chemical

Agency’s ECHA CHEM database, as well as a derived no-effect level (DNEL) for oral

exposure.

12

2 ANALYTICAL METHOD DEVELOPMENT FOR BFRs IN HOUSE DUST

The principle aim of this method development is to develop and validate an effective analysis

method for indoor dust containing selected novel and emerging BFR, OPFR, as well as

PBDEs. These compounds are analyzed together in the same sample. The aim of method

development is to achieve a reasonable extraction solvent volume, small sample amount

consumption, simple and easy clean-up, minimal loss of sample, technical feasibility with

available machines and techniques, short extraction time and high sample throughput.

The analytical method used throughout the method development was based on Van den Eede

et al. (2012). Gas Chromatography - Electron Impact – High Resolution Mass Spectrometry

(GC-EI-HRMS) was used to analyze the samples. The method follows the following general

protocol described in section 2.1 below.

Between May-August 2015, three development tests were done. The tests performed were a

continuation of the development tests that had been performed at THL before May 2015. The

three tests done were coded 15T024, 15T029 and 15T031.

2.1 MATERIALS AND METHODS

2.1.1 CHEMICALS AND INSTRUMENTS

Samples Used

During the course of the method development, samples used include a prepared 12C mixture,

Standard Reference Material 2585 of indoor dust, and actual samples of air filter, settled dust,

and air conditioner filter dust. In the validation test, high and low-level spiked samples were

also used as a comparison to actual samples. Typically, 50 mg of dust sample was weighed in

each test tube for extraction. One hundred μL 13C Internal Standards (2.5-6.25 ng/sample in

toluene) were added to the sample before extraction.

Florisil Clean-Up Columns

Prepared activated Florisil cleanup columns in 3 ml glass tubes were used for the clean-up

process. Florisil was activated before use in a 200°C oven overnight and cooled in a

desiccator before use. Three ml:n LiChrolut SPE tubes, of 6 cm in length, by Merck were

13

used. A small piece of cotton wool was fitted to the bottom of the tubes. Columns were

packed with 2cm layer of activated Florisil. A small cotton wool plug was added on top of

each column to prevent dusting of the Florisil when solvent was poured on top. Florisil

columns were washed with 2.5*2 ml 50% dichloromethane-hexane (dcm-hex) before

fractionation.

H2SO4-Silica Clean-up Columns

Prepared H2SO4-silica clean-up columns were subsequently added under Florisil column for

Fraction 1 only. Three ml LiChrolut SPE tubes, of 6 cm in length, by Merck were used. A

small piece of cotton wool was fitted to the bottom of the tubes. Columns were packed with 2

cm layer of 44% volume-per-weight H2SO4-silica. A small cotton wool plug was added on top

of Florisil column to prevent dusting when solvent was poured on top. H2SO4-silica columns

were washed with 2.5*2 ml 50% dcm-hex before fractionation. In the sections that follow, test

procedures are described backwards (GC-HRMS, fractionation and cleanup, extraction) as

compared to order of sample preparation (extraction, fractionation and cleanup, GC-HRMS)

because GC-HRMS method has to be in place before anything else can be tested. However,

there is naturally overlap and feedbacks between all of these phases.

2.1.2 GC-HRMS CONDITIONS

GC-MS analysis was performed with an HP 6890 gas chromatograph (Agilent, Little Falls,

DE, USA) coupled to an Autospec Ultima high resolution mass spectrometer (Waters,

Manchester, GB). The system was equipped with PTV injector and CTC CombiPal

autosampler. MassLynx 4.0 software was used for instrument control and data analysis.

Analysis of fraction 1 containing selected BFRs was as follows: empty multi baffle liner

(Agilent, Part# 5183-2037) was used in the PTV inlet. A Phenomenex ZB-5MSplus (Part No.

7FD-G030-08) capillary column cut to a length of 6 m, 0.18 mm I.D., and 0.18 µm film

thickness was used. The injection volume was 2 µl. The program for PTV splitless injection

was: 90 °C for 0.10 min, 600 °C/min to 300 °C, hold 1.0 min, 700 °C/min to 325°C, hold 7.0

min. Vent time was 0.09 min, vent flow 80 ml/min and vent pressure 100 kPa. Purge time was

1.25 min. Constant helium gas flow was 1.0 ml/min. Oven temperature program was: 70 °C

for 1.25 min, 20°C/min to 240°C, 50°C/min to 300 °C, 15 °C/min to 320 °C, hold 12.0 min.

14

For PFRs GC-HRMS analysis was as follows: a single baffle liner (Zebron, Part No: AG2-

1F06-05) with deactivated cotton wool plug in the bottom was used in the PTV inlet. An

Agilent DB-5MS UI (Part No. 122-5532UI) capillary column of 30 m, 0.25 mm I.D., and 0.25

µm film thickness was used. The injection volume was 1 µl. The program for PTV splitless

injection was: 80 °C for 0.10 min, 720 °C/min to 280 °C, hold 2.0 min, 720 °C/min to 320 °C,

hold 20.0 min. Vent time was 0.09 min, vent flow 100 ml/min and vent pressure 80 kPa.

Purge time was 2.00 min. Constant helium gas flow was 1.0 ml/min. Oven temperature

program was: 70 °C for 1.00 min, 15 °C/min to 150 °C, 10 °C/min to 280 °C, 7 °C/min to

320 °C, hold 12.0 min.

MS-parameters for both BFRs and PFRs were: MS transfer line temperature 300 °C,

temperature of ionization chamber 280 °C and energy of EI+ ionization 35 eV. The two most

intense and/or interference free ions of each compound were selected for monitoring. Sample

peak identification was based on the matching of retention times and ion ratios with those of

standards.

2.1.3 FRACTIONATION AND CLEAN-UP

Determination of Suitable Eluent

Test 15T024 was conducted on 12 May 2015. The aim of this test was to separate BFR and

OPFR each to their own fraction. This test endeavoured to determine the weakest eluent

possible to elute all OPFRs from the Florisil column. Weakest eluent implies that the elution

of impurities can be minimized. This test was performed with Florisil column only. 12-C

standards of known BFR and PFR were used for the elution test. One hundred μl of 12-C

standards, containing 10-25ng of target compounds was used in each sample. Target

compounds were added to the columns in 500μl of hexane and 1 drop of nonane. No dust

material was analyzed and therefore no extraction was performed. After fractionation, 100μl

of 13-C internal standards were added to each 12-C mixture fraction. Samples were then

concentrated and analyzed by GC-EI-MS. Table 2 shows the 12C standard mixture that was

used in this test. Table 3 shows the internal standard and recovery standard used in this test

and the subsequent tests.

15

Table 2. 12C Mixture used in Test 15T024.

16

Table 3. Internal and Recovery Standard used in Test 15T024 and subsequent tests.

17

2-Step Clean-Up

A 2-step clean-up was then performed, resulting in fractionation of analytes and removal of

interferences. Dual clean-up columns, with Florisil column on top of H2SO4-silica column

was used for Fraction 1 only. To elute Fraction 1 containing BFR (except BEH-TEBP),

extracts were added to dual-column according to the columns’ capacity, taking care not to

overflow. Samples were then eluted with 2*2.5ml of eluent 1 until all solvent had come

through both Florisil and H2SO4-silica columns. Each sample was collected in a 10ml test

tube. For Fraction 2, containing OPFR, the H2SO4-silica columns were removed, and eluate

from Florisil column was collected separately in a clean 10ml test tube. Florisil columns were

eluted with 2*2.5ml of eluent 2 until all solvent has come through.

Fraction 1 contains most of the selected BFR. Fraction 2 contains most of the selected OPFR.

The aim of the clean-up was to have most BFRs in Fraction 1 and most OPFRs in Fraction 2.

Fig. 1 Dual clean-up columns for Fraction 1, Florisil column on top of H2SO4-silica column.

Figure 1 is a photograph of the dual clean-up columns for Fraction 1, with Florisil column on

top of H2SO4-silica column.

18

Determination of Best Clean-Up Technique

Test 15T031 was conducted on 16 June 2015. This test attempted to extract selected BFRs

and OPFRs from Standard Reference Material of Indoor Dust SRM 2585. Extraction was

performed as described in Section 2.0 with 2ml of dichloromethane as the extraction solvent.

Fractionation was performed with a dual-column set-up as described above and shown in Fig.

1.

The aim of this test was to test three different fractionation techniques for the best clean-up

result and method simplicity. Each technique to be tested was added to two samples. The

following fractionation techniques were tested:

1. Direct pouring of combined 2+2ml dichloromethane extract to dual fractionation-

clean-up columns.

2. Evaporation of combined 2+2ml dichloromethane extract to 1ml dichloromethane

before adding to dual-columns.

3. Evaporation of combined 2+2ml dichloromethane extract to 0.5ml dichloromethane,

and dilution with 0.5ml hexane before adding to dual-columns.

Dual clean-up columns, with Florisil column on top of H2SO4-silica column were used for

Fraction 1 only. For Fraction 1, containing BFRs, extracts were added to each dual-column

according to the columns’ capacity, taking care not to overflow. Samples added with above-

listed techniques 1 and 2 were eluted with 2*2.5ml of dichloromethane until all solvent has

come through both Florisil and H2SO4-silica columns. Samples added with above-listed

technique 3 were eluted with 2*2.5ml 50% dichloromethane-hexane. Each sample was

collected in a 10ml test tube.

For Fraction 2, containing OPFR, the H2SO4-silica columns were removed, and elute from

each column was collected separately in a clean 10ml test tube. Florisil columns were eluted

with 2*2.5ml of 10% acetone-dichloromethane until all solvent has come through.

After fractionation and clean-up, 30μL of nonane was added to each sample. Elutes were then

concentrated and analyzed as described in this section.

19

Na2SO4 clean-up of Fraction 2

In Test 15T029, which will be elaborated in Section 2.1.4, interferences in the MS

chromatograms were observed in Fraction 2. Eluates for Fraction 2 for Test 15T029, which

had also been concentrated and transferred to autosampler vials, went through a further clean-

up step with Na2SO4. One ml of hexane was first added to each autosampler vial containing

FR compounds in 120μL of toluene and 30μL of nonane. The contents of each vial were then

transferred to a 5ml test tube with a Pasteur pipette. Each autosampler vial was then flushed

with a further 1.0ml of hexane, which was also transferred to the corresponding test tube.

Approximately 500μg of activated Na2SO4 was added to each test tube. Test tubes were then

shaken in the VWR-shaker for 4 minutes at 1800rpm in pulse mode (3 seconds on, 1 second

off). At this point, the yellow colour was reduced but not completely removed. Samples were

centrifuged in 3000rpm for 2*2 minutes. The hexane (containing FR) in each test tube was

then decanted to a clean 5ml test tube. Shaking with VWR and centrifuging were then

repeated on the sample containing Na2SO4. The two portions of hexane from each sample

were then combined in the same clean test tube.

Concentration and GC-MS Analysis

After fractionation and clean-up, 30μL of nonane was added to each sample. Elutes were

concentrated under a gentle stream of nitrogen gas to a volume of about 500μL before transfer

to GC-MS autosampler vials. Each test tube was flushed with 2*200μL of dichloromethane to

ensure complete transfer of target compounds to autosampler vial. Fifty μL of 13C Recovery

Standards (5 – 10 ng/sample in hexane) were added before GC-MS analysis. Thirty μL of

nonane was also added to each autosampler vial. (No additional nonane would be added if

nonane had already been added to samples before evaporation.) Vials were covered with

wadding and evaporated in fume hood overnight. Cleaned and concentrated extracts were then

analyzed in mixture of 120μL of toluene and 30μL of nonane.

20

2.1.4 EXTRACTION

Extraction of FR from Dust Sample

Extraction was done by putting the sample at 1800 rpm to VWR Shaker for 4 minutes in a

pulse mode of 3 seconds on, 1 second off, followed by 3500 rpm centrifugation for 2 min.

Two ml of extraction solvent was used for each dust sample. Extraction was then repeated

with a further 2ml of extraction solvent. 2+2 ml of extract for each sample were then

combined.

During the method development, different extraction and elution solvents were tested, namely

hexane, dichloromethane, acetone, diethylether and their appropriate mixtures.

Determination of Suitable Extraction Solvent

Test 15T029 was conducted on 26 May 2015. This test attempted to extract selected BFRs

and OPFRs from the intended medium. Standard Reference Material of Indoor Dust SRM

2585 was used as samples for this test. The aim of this test was to determine a suitable

extraction solvent. Three different solvents: 100% Dichloromethane, 25% Acetone-Hexane,

50% Dichloromethane - Ether were tested in this test. Both 100% Dichloromethane and 25%

Acetone-Hexane were determined to be suitable extraction solvents for polar BFRs and

OPFRs by Van den Eede et al. (2012). 50% Dichloromethane-Ether was a new solvent to be

tested. It was noted that the ether had to be carefully evaporated away for the H2SO4-silica

clean-up applied to the OPFR fraction.

2.1.5 VALIDATION

A validation was subsequently performed for the method development. This validation

evaluates the performance of the analysis of brominated compounds. Performance of the

analysis of organophosphorous compounds is also evaluated, and possible further

developmental needs identified, especially in sample clean-up.

Table 4. shows the type and description of the samples used for the validation test. Samples

were prepared and analysed according to the method previously described.

21

Table 4. Type and description of the samples used for the validation test.

Blank No sample. Only extraction solvent 100%

Dichloromethane

National Institute of Standards and Technology (NIST)

SRM-2585

Standard Reference Material – 2585 House

Dust

Home exhaust air filter dust from a private home (Home 1) Indoor dust collected from exhaust air filter

with a vacuum cleaner. Dust accumulated

through several months. Collected on 6

January 2015 and 24 July 2015 and mixed.

Home settled dust from a private home (Home 1) Indoor dust collected from home surfaces

with a vacuum cleaner. Collected on 25 July

2015.

Home air condition filter dust

rom a private home (Home 2)

Indoor dust collected from air condition

filter dust with vacuum cleaner. Collected

on 29 July 2015.

Low-level spiked home exhaust air filter dust Home exhaust air filter dust spiked

High-level spiked home exhaust air filter dust Home exhaust air filter dust spiked

Preparation of Spiking Solution and Spiking of House Dust

Spiked house dust is prepared from dust collected from home exhaust air filter dust of a

private home (described above).

100% dichloromethane was used as spiking solvent. Approximately 6ml of spiking solvent

and an appropriate volume of BFR stock solution was added to 1.0g of dust in an Erlenmayer

flask and mixed in VWR-shaker at 500rpm for 30 minutes. The flask was uncapped and

solvent let evaporate overnight at room temperature in fume hood, until the original weight of

the dust sample had been achieved. The spiked dust was then re-homogenized by shaking in

VWR-shaker for 3 minutes at 2500 rpm. One g of low level spiked sample, and another 1.0g

of high level spiked sample were prepared. Tables 5 and 6 below show the concentration of

each compound in the stock solution and the final spiked concentration to the dust.

22

Table 5. FR concentration in the stock solution and concentration of FR spiked to the low

level spiked sample.

Compound

Concentration in

the stock solution

(ng/ml)

Pipet to test

tube containing

dcm (µl)

Mass in

test tube

(ng)

Concentration

spiked to the

dust (ng/g)

TCEP 100

100

10 10

TCIPP 100 10 10

TDCIPP 100 10 10

TDBPP 100 10 10

TiBP 100 10 10

TnBP 100 10 10

TBOEP 250 25 25

EHDPP 100 10 10

TEHP 100 10 10

TPHP 100 10 10

TMPP 100 10 10

ab-DBE-

DBCH 200 20 20

PBT 100 10 10

TBP-DBPE 100 10 10

EH-TBB 250 25 25

BTBPE 100 10 10

BEH-TEBP 100 10 10

DBDPE 250 25 25

BDE-28 50

100

10 10

BDE-47 50 10 10

BDE-99 50 10 10

BDE-100 50 10 10

BDE-153 35 7 7

BDE-154 50 10 10

BDE-183 50 10 10

BDE-209 160 250 40 40

23

Table 6. FR concentration in the stock solution and final concentration of FR spiked to the

high level spiked sample.

Compound

Concentration in

the stock solution

(ng/ml)

Pipet to test

tube containing

dcm (µl)

Mass in

test tube

(ng)

Concentration

spiked to the

dust (ng/g)

TCEP 100

1000

100 100

TCIPP 100 100 100

TDCIPP 100 100 100

TDBPP 100 100 100

TiBP 100 100 100

TnBP 100 100 100

TBOEP 250 250 250

EHDPP 100 100 100

TEHP 100 100 100

TPHP 100 100 100

TMPP 100 100 100

ab-DBE-DBCH 200 200 200

PBT 100 100 100

TBP-DBPE 100 100 100

EH-TBB 250 250 250

BTBPE 100 100 100

BEH-TEBP 100 100 100

DBDPE 250 250 250

BDE-28 50

1000

100 100

BDE-47 50 100 100

BDE-99 50 100 100

BDE-100 50 100 100

BDE-153 35 70 70

BDE-154 50 100 100

BDE-183 50 100 100

BDE-209 160 2500 400 400

24

In the validation, the following parameters were determined:

• Limit of Detection (LoD) = 3*SD of blank sample

• Limit of Quantification (LoQ) = 8*SD of blank sample

• Accuracy (as compared with reference values and other labs)

• Precision (relative standard deviation for each compound)

• Measurement Uncertainty (based on accuracy and precision)

• Suitability of chromatographic system

Determination of Limit of Detection (LOD) and Limit of Quantification (LOQ)

Limit of detection and limit of quantification were calculated as follows:

LOD = 3 * SD

LOQ = 8 * SD

SD is the standard deviation of the blank samples producing detectable signal with correct ion

ratios, i.e. +/- 20% of which in calibration samples or theoretical values. If no signal from

blank sample is obtained, LOD and LOQ were calculated as 95th percentile of the

concentrations corresponding to signal to noise ratios – 3:1 for LOD and 8:1 for LOQ,

produced by MassLynx software from all actual samples.

Accuracy and Precision

Accuracy was determined as the percent deviation of the analysis results SRM-2585 from the

certified values. Comparison was more qualitative for compounds with only reference values

from other laboratories. Percent recovery from spiked samples was also used to estimate

accuracy. Precision was calculated as the relative standard deviation of the results from all

dust samples.

Accuracy -1: For 2 batches of actual samples, 4 parallel samples of SRM-2585 were measured

and their results compared to certified or referenced values.

Accuracy – 2: For 2 batches of samples, 4 parallel samples of spiked dust were measured and

percentage recovery was calculated for all compounds as follows:

25

% Recovery = 100*(Average results of spiked samples – Average of non-spiked results)/

Spiked concentration

Precision: Relative standard deviation was calculated separately from parallel samples for

each different house dust sample. Combined precision was calculated for each compound as

the root mean square of Relative Standard Deviation - % from all different dust samples. If

large deviations in the range of concentrations occur, precision can be calculated for different

ranges of concentrations.

Measurement Uncertainty

Measurement uncertainty was calculated as a combination of accuracy and precision.

Measurement Uncertainty: (utot)2=(uAccu)2+(uPres)2

Expanded Measurement Uncertainty with 95 % confidence interval

MU=2*utot

utot = Total MU

uAccu = Accuracy

uPres = Precision

Suitability of the Chromatographic System

This validation aimed to have chromatographic peaks that are symmetrical and without

interfering extra peaks in the channel of the compound to be measured. Also, there was to be

no serious interferences in the HRMS lock masses at the retention times of compounds to be

measured. Recovery rates of internal standards would preferably be in the range between 60%

and 120%. Lower recovery rates would be acceptable for highly volatile compounds. Ion

ratios of compounds to be measured should be within +/- 20% of the theoretical values or

values obtained from calibration standards.

26

2.2 RESULTS AND DISCUSSION

2.2.1 FRACTIONATION AND CLEAN-UP OPTIMIZATION

Results for Test 15T024

In this test, the standard 12C mixture of compounds in 500 μl of hexane were added to cleanup

columns. It was concluded that 100% Dcm was the most suitable solvent for the elution of

Fraction 1 from the Florisil column. With this solvent most of the BFRs could be separated

from the OPFRs. Ten% Acetone-Dcm was determined to be the most suitable solvent for the

elution of Fraction 2, where all of OPFR, could be eluted, except that 50% of TBOEP still

remained in the column. However, the use of 13C-labelled internal standard for TBOEP

corrects for this loss. BEH-TEBP was eluted in Fraction 2 together with OPFR, and would

remain so in subsequent tests.


In this test it was found that the detected concentration of each compound was quite consistent

across all 3 treatments. Table 7 below illustrates the results for this test.

Table 7. The detected concentration of each compound was quite consistent across all 3

treatments. BFR Extract (4 ml of

DCM) directly

to dual-column

Extract evaporated

to 1mL then to

dual-column

Extract evaporated to

0.5ML, 0.5mL hex added

then to dual-column

Certified/reference

values (range)

ng/g

Average Average 15T029-6-F1

ng/g ng/g ng/g

ab-DBE-DBCH 6 3 3

PBT 0 0 0

TBP-DBPE 0 0 0

EH-TBB 17 20 21 26-40 (2-6)

BTBPE 15 14 20 32-76 (4-14)

BEH-TEBP 1898* 6216* 0* 145-1300(16.7-

94)*

DBDPE 0 0 0 <20

BDE-28 24 24 25 46.9 (4.4)

BDE-47 243 249 279 497 (46)

BDE-100 74 78 95 145 (11)

BDE-99 486 502 577 892 (53)

BDE-154 40 41 49 83.5 (2.0)

BDE-153 59 60 68 119 (1)

BDE-183 no data no data no data 43.0 (3.5)

BDE-209 2306 2241 2336 2510 (190)

* It was found that BEH-TEBP had degraded in H2SO4-silica , therefore, these results for BEH-TEBP for Fraction 1 were not reliable.

27

The average concentration of compounds for 15T031 detected was approximately 50% of

certified/reference values. This error was due to calibration and was corrected in subsequent

tests. However, in the interest of determining the most simple and suitable fractionation

technique, the above technical issues were not pursued further.

In Test 15T029, activated Na2SO4 was found to be ineffective in removing impurities in

Fraction 2. Tests for the cleanup of Fraction 2 were not continued further.

2.2.2 EXTRACTION


The above-described clean-up facilitated the GC-HRMS analysis of Fractions 1-A and 1-B.

Based on the finding that the detected concentration of BFR and PBDE were consistent across

all three treatments in Test 15T031, direct pouring of 4ml Dcm extract was chosen for future

implementation. As Table 8 shows, the concentrations for PBDEs analyzed from SRM2585

by different solvents was generally consistent with certified values.

Table 8. The concentrations for PBDEs analyzed from SRM2585 by different extraction

solvents. Certified Value (SRM-

2585)

100%

Dichloromethane

25% Acetone-

Hexane

50%

Dichloromethane

-Ether

ng/g Average (ng/g) Average (ng/g) Average (ng/g)

BDE-28 46.9 (4.4) 45 46 45

BDE-47 497 (46) 531 519 524

BDE-100 145 (11) 175 170 173

BDE-99 892 (53) 1019 1009 1034

BDE-154 83.5 (2.0) 83 81 83

BDE-153 119 (1) 114 111 112

BDE-183 43.0 (3.5) 42 42 36

BDE-209 2510 (190) 2570 2871 2549

All three of the elution solvents tested, namely, 100% Dichloromethane, 25% Acetone-

Hexane, 50% Dichloromethane - Ether were shown to be equally efficient in extraction of

BFR. Table 9 shows that the amount of BFRs extracted are comparable across all three

elution solvents.

28

Table 9. The amount of BFRs extracted are comparable across all three elution solvents.

BFR Comparison value

(range)

100%

Dichloromethane

25%

Acetone-

Hexane

50%

Dichlorometh

ane-Ether

Average (ng/g) Average

(ng/g)

Average (ng/g)

ab-DBE-

DBCH

5 3 3

PBT 0 0 0

TBP-DBPE 0 0 0

EH-TBB 26-40 (2-6) 38 38 39

BTBPE 32-76 (4-14) 50 33 33

BEH-TEBP 145-1300(16.7-94) n/a n/a n/a

DBDPE <20 0 1 1

100% dichloromethane was chosen to be the most suitable extraction solvent for subsequent

extractions due to ease of preparation and use. It was found that an additional H2SO4- silica

clean-up column was necessary for the BFR fraction in order for the extract to be sufficiently

clean for the GC-HRMS analysis. The main benefit of using 100% dichloromethane is that it

can be directly poured to dual Florisil - H2SO4- silica clean-up column after extraction

without prior evaporation that is needed for 25% Acetone-Hexane and 50% Dichloromethane-

Ether.

29

2.2.3 SUMMARY OF SAMPLE TREATMENT IN FLOW CHART

Fig. 2 Flow-chart of analysis method.

Figure 2 shows a flow-chart of the analysis method.

2.2.4 RESULTS OF VALIDATION

Summary of Results

LOQ & MU were acceptable for all BFRs. LOQ of BFRs ranged between 0.5 – 5.0 ng/g. For

OPFR, LOQ ranged between 6.9-613 ng/g. Precision for OPFR was good, meaning the

relative SD were consistent for each compound. Accuracy for OPFR was poor compared to

other labs’ analysis result for the certified material for indoor dust SRM 2585. Therefore,

method for OPFR analysis requires further testing that was not conducted within the scope of

this thesis. Tables 10, 11, 12 and 13 below present a summary of validation results, taken

from an internal validation report written by Panu Rantakokko, PhD, Senior Researcher of

THL on 11 September 2015.

30

Table 10. Summary of LODs, LOQs and MUs to be reported for BFRs.

Compound LOD (ng/g) LOQ (ng/g)

MU (%)

< 50 ng/g

MU (%)

> 50 ng/g

ab-DBE-DBCH 1.0 2.5 55 55

PBT 0.2 0.5 55 40

TBP-DBPE 0.2 0.5 65 45

EH-TBB 2.0 5.0 25 25

BTBPE 0.2 0.5 25 25

BEH-TEBP* 10 25 100 50

DBDPE 2.0 5.0 30 30

BDE-28 0.2 0.5 25 20

BDE-47 0.2 0.5 30 20

BDE-100 1.0 2.5 40 40

BDE-99 1.0 2.5 25 25

BDE-154 0.3 0.7 25 25

BDE-153 0.3 0.7 30 30

BDE-183 0.6 1.5 35 35

BDE-209 1.0 2.5 75 75

Table 11. LODs and LOQs based on blank (n=4) and MassLynx (OPFRs, n=21).

Compound

/Sample LOD (ng/g) LOQ (ng/g)

TIBP 20 52

TNBP 12 32

TCEP 25 67

TCIPP 230 613

TDCIPP 21 57

TPHP 2.6 6.9

TBOEP 6.5 17

EHDPP 34 91

TEHP* 40 106

TMPP 7.1 19

*TEHP has very poor sensitivity with the ions selected, but alternative ions would be low

mass and extremely noisy and non-specific.

31

Table 12. Accuracy, precision and MU from results of SRM 2585 a (BFRs, n=8).

COMPOUND

Average (range)

(ng/g)

RSD

(%)

Certified/Other

(ng/g)

Recovery

(%) MU (%) Source

ab-DBE-DBCH <LOQ

PBT 0.22 (0.12-0.26) 24

TBP-DBPE <LOQ

EH-TBB 33 (31-36) 5.0 26 127 56 Van den Eede 2012

BTBPE 54 (35-86) 40 b 39 140 b 112 b Van den Eede 2012

BEH-TEBP c 1212 (1141-1281) 5.6 574 211 c 223 c Van den Eede 2012

DBDPE <LOQ <7.1 Van den Eede 2012

BDE-28 49 (47-51) 3.5 46.9 104 11 Certified

BDE-47 487 (443-551) 8.2 497 98 17 Certified

BDE-100 164 (148-191) 9.5 145 113 32 Certified

BDE-99 954 (870-1114) 8.5 892 107 22 Certified

BDE-154 83 (78-96) 7.6 84 99 15 Certified

BDE-153 115 (109-133) 7.5 119 96 17 Certified

BDE-183 37 (33-46) 11 43 87 34 Certified

BDE-209 3138 (2368-4807) 26 d 2510 125 d 72 d Certified a SRM 2585 has certified concentrations for PBDEs only. For other BFRs many sources exist, but Van den

Eede represents generally a recognized high quality laboratory. b For BTBPE some results were outliers, especially in the series 2. Source of deviation needs to be tested in

future work. c Sahlström et al 2012 measured a concentration of 1300 ng/g for BEH-TEBP. d Range of results for BDE-209 is large. Possible laboratory contamination needs to be tackled in future

testing.

Table 13. Accuracy, precision and MU from results of SRM 2585 a (OPFRs, n=4).

COMPOUND

Average

(range) (ng/g)

RSD

(%) b

Certified/

Other (ng/g)

Recovery

(%) b

MU

(%) Source

TIBP 18 101.5

TNBP 1196 1.6 190 629 1059 Van den Eede 2012

TCEP 3363 6.2 680 495 789 Van den Eede 2012

TCIPP 3780 9.8 860 440 679 Van den Eede 2012

TDCIPP 7744 14.0 3180 244 288 Van den Eede 2012

TPHP 4335 5.9 1160 374 548 Van den Eede 2012

TBOEP 57568 7.5 63000 91 23 Van den Eede 2012

EHDPP 3039 9.5 1300 234 268 Bergh 2012

TEHP 1091 9.1 370 295 390 Bergh 2012

TMPP 3435 10.0 1140 301 403 Van den Eede 2012

32

Suitability of Chromatographic System

For the BFRs, percentage recovery of internal standards were found to be in the range of 60%

- 120%. Ion ratios of compounds to be measured were within +/- 20% of the theoretical

values, or values from calibration standards.

For the OPFRs, peak sizes tended to be too small in the calibration standard for all

compounds, and for some compounds in the Internal Standard solution added to samples.

Table 14 below illustrates the amount of dust detected in Home Settled Dust samples (n=4),

Home Exhaust Air Filter Dust samples (n=4) and Home Air Condition Filter Dust samples

(n=4) during the validation process. Home settle dust samples and home exhaust air filter dust

samples are from the same home (Home 1), while Home air condition filter dust samples are

from another home (Home 2).

33

Table 14. Amount of dust detected in Home Settled Dust samples, Home Exhaust Air Filter

Dust samples, and Home Air Condition Filter Dust samples during validation. Vapor pressure

of each compound is included for comparison. Sample

type

Settled Dust (n=4,

Home 1)

Exhaust Air Filter Dust

(n=4, Home 1)

Air Condition Filter Dust

(n=4, Home 2)

Compound Average

(ng/g)

RSD

(%)

Average (ng/g) RSD

(%)

Average (ng/g) RSD

(%)

Vapour

Pressure

(Pa)(25C)

ab-DBE-

DBCH

0.56 29.2 0.41 22 0.31 5.8 2.97E-03

PBT 0.70 11 0.63 36a 1 19 6.00E-04

TBP-

DBPE

0.64 4.9 2.0 5.4 0.08 13 1.26E-05

EH-TBB 5.6 15 8.7 3.4 2.7 23 3.71E-07

BTBPE 3.5 6.1 2.7 9.2 8.5 135 3.88E-10

BEH-

TEBP

136 19 138 8.2 192 11 1.55E-11

DBDPE 158 17 212 5.7 327 140 n/a

BDE-28 0.95 5.1 1.2 6 0.27 8.1 n/a

BDE-47 23 4.3 24 2.7 4.1 4.1 n/a

BDE-100 4.6 3.9 4.5 10.7 0.94 7.7 n/a

BDE-99 41 2.9 38 2 5.3 11 n/a

BDE-154 2.4 5.6 2.7 1.9 0.49 31 n/a

BDE-153 5.6 7.2 8.0 3.9 1.2 60 n/a

BDE-183 1.3 14 2.2 37 2.9 98 n/a

BDE-209 752 2.2 785 3.6 184 36 n/a

34

2.2.5 CONCLUSIONS OF METHOD DEVELOPMENT AS OF 2016

It was concluded that 100% Dcm was the most suitable solvent for the elution of Fraction 1

from the Florisil column. With this solvent most of the BFRs could be separated from the

OPFRs. 10% Acetone-Dcm was determined to be the most suitable solvent for the elution of

Fraction 2, where all of OPFR, could be eluted, except that 50% of TBOEP still remained in

the column. BEH-TEBP was eluted in Fraction 2 together with OPFR. Direct pouring of 4ml

Dcm extract was chosen for future implementation. 100% dichloromethane was chosen to be

the most suitable extraction solvent for subsequent extractions due to ease of preparation and

use. Settled dust was considered to be a preferable matrix.



values, or values from calibration standards. For the OPFRs, chromatography peak sizes

tended to be too small in the calibration standard for all compounds, and for some compounds

in the Internal Standard solution added to samples.

For the validation, Limit of Quantification (LOQ) & Measurement Uncertainties were

acceptable for all BFRs. LOQ of BFRs ranged between 0.5 – 5.0 ng/g. For OPFR, LOQ

ranged between 6.9-613 ng/g. Precision for OPFR was good, but accuracy was poor compared

to other labs’ analysis result for the certified material for indoor dust SRM 2585. Therefore,


this thesis.

As compared to sampling of settled dust by collecting settled dust on surfaces with a vacuum

cleaner, indoor dust collected on the exhaust air filter could be a good time and space

integrated sample from the entire indoor space of a household, in the case where this

particular exhaust mechanism is installed. Therefore, exhaust filter dust can be representative

of one indoor compartment as all air that exit the house goes through the filter. However, the

concentration of more volatile BFRs such as ab-DBE-DBCH and PBT tend to be lower than

that on settled dust, as illustrated in Table 14. Therefore, settled dust would be a preferable

matrix than exhaust dust within the scope of this particular validation test described in this

thesis. Air conditioner filter dust showed large deviation, as illustrated by the high relative

standard deviation in Table 14. Therefore, air conditioner filter dust was not an ideal matrix.

35

Figure 3 shows a schematic of an example of a heat recovery unit of a house ventilation

system with Stale air from inside filter used for sample collection.

Figure 3. Schematic illustration of exhaust ventilation. This figure has been provided by Dr.

Panu Rantakokko of the National Institute for Health and Welfare (THL), Kuopio.

It was noticed that the spiked samples were more homogenous than non-spiked samples when

1g of sample was spiked at one time. The reason may be that the FRs in the samples have

been extracted and equally redistributed during the spiking process with dichloromethane,

resulting in a more homogeneous overall distribution of the FRs present in the sample.

However, it would be difficult to obtain such a large quantity of indoor dust sample from a

single site unless taken from exhaust air filter. Large dust mass available from the filter is one

significant benefit of using it for dust sampling.

2.2.7 DISCUSSION

It was recommended that elution of BEH-TEBP need to be tested further to reduce BEH-

TEBP degradation in H2SO4-silica clean-up column. There was an aim to get BEH-TEBP to

36

Fraction 1. However, it has since proven to be unsuccessful. OPFR accuracy needs to be

improved, but at the present, difficulties with impurities remain. There may be a need to

increase OPFR concentrations in Internal Standard and Calibration Standard Solutions to

match the levels found in actual samples. Also, it may be good to have separate calibration

solutions for BFR and OPFR.

3. HUMAN HEALTH RISK ASSESSMENT

According to the definition put forward by the United States Environmental Protection

Agency (USEPA 2017), human health risk assessment is a process of estimating the

probability of human health effects, especially adverse health effects for a given exposure to a

substance, which can be a chemical present in an environmental medium. There are four basic

steps to human health risk assessment: hazard identification, dose-response assessment,

exposure assessment and risk characterization. Hazard identification attempts to determine

whether a substance may cause harm to humans by putting together available information on

toxicokinetics and possible adverse effects of a chemical on human health. Dose-response

assessment attempts to determine a numerical relationship between exposure to the substance

and the effects. The dose-response relationship links the probability and severity of adverse

health effects to the level of exposure to the substance. Exposure assessment is a process of

determining the magnitude of exposure, frequency of exposure and duration of exposure to

the substance. Exposure assessment also takes into consideration the population exposed to

the substance. Risk characterization summarises the information gathered for the first three

steps of the risk assessment process, and from these information, conclusions may be drawn

regarding the extent of risk resulting from exposure to a substance.

Figure 4 below shows a schematic representation of the four steps to human health risk

assessment. This figure was taken from the USEPA website (USEPA 2017).

37

Figure 4. The four-step process to human health risk assessment.

Hazard characterization in this section will be based on currently available information from

literature. The endpoint Derived No-Effect Level (DNEL) will be employed. Risk

Characterisation Ratio will be used to perform hazard characterization.

The DNEL is a part of human health hazard assessment stipulated in Annex I of REACH,

which comprises of four steps: evaluation of non-human information, evaluation of human

information, classification and labelling, and finally, the derivation of DNEL (Munn 2007).

DNEL is the level of human exposure that should not be exceeded. In hazard characterization,

the estimated exposure of a population is compared with the corresponding DNEL. The risk

of adverse health effect is considered to be adequately controlled if the exposure level does

not exceed the corresponding DNEL, as stipulated in REACH Annex I, Section 6.4 (ECHA

2016a).

The Risk Characterisation Ratio (RCR) is a way to quantify risk in a given exposure scenario.

Based on the guidance from ECHA (2016a), the RCR is calculated as follows:

RCR = Exposure/DNEL

If exposure is less than DNEL, i.e. RCR<1, the risk is adequately controlled. If exposure is

larger than DNEL, i.e. RCR>1, the risk is not adequately controlled.

38

This section will focus on the six novel and emerging BFR listed in Table 1, namely, ab-

DBE-DBCH, PBT, TBP-DBPE, EH-TBB, BTBPE and BEH-TEBP. The OECD eChem

portal, HSDB database and the ECHA CHEM database were consulted for available relevant

information on the above substances.

There was insufficient toxicological information available for ab-DBE-DBCH, PBT, TBP-

DBPE to perform a hazard identification. There is some information available for BTBPE

from the Scientific Opinion on Emerging and Novel Brominated Flame Retardants (BFRs) in

Food, issued by the European Food Safety Authority in 2012. An evaluation of human-health

related toxicity has been included in the United States Environmental Protection Agency

Alternatives Assessment update (2015) for EH-TBB and BEH-TEBP. However, the substance

EH-TBB is not found on the OECD eChem portal. Chemical registration dossier (ECHA

2016b) is available for BEH-TEBP through the European Chemical Agency’s ECHA CHEM

database. In this dossier, a Derived no-effect level (DNEL) for oral exposure has been

estimated for BEH-TEBP.

This section summarises relevant toxicological information available for the substances

BTBPE, EH-TBB and BEH-TEBP. An exposure estimation has been performed based on the

amount of BFR detected from a recent unpublished study by THL. Based on this exposure

estimation, Risk Characterisation Ratio based on the Guidance on Information Requirements

and Chemical Safety Assessment (ECHA 2016a) has been calculated for BEH-TEBP

according to the DNEL for oral exposure.

The substance Bis(2-ethyl-1hexyl)tetrabromophthalate (BEH-TEBP) with CAS Number

26040-51-7 was selected for hazard characterization based on the availability of a DNEL

value for oral exposure for the general population.

The DNEL value for BEH-TEBP has been derived for long-term oral exposure according to a

repeated dose oral toxicity study which will be described in Section 3.3.2.

A commercial mixture FM-550 containing EH-TBB and BEH-TEBP has been used in several

toxicological experiments mentioned in this section. The exact composition of FM-550 is

39

proprietary. However, it has been found by Stapleton et al. (2008) that FM-550 contains

approximately 50% of isopropylated triaryl phosphate and triphenylphosphate. The other 50%

consisted of brominated compounds EH-TBB and BEH-TEBP in approximately 4:1 ratio by

mass.

Another commercial mixture, FMBZ-54 has also been used in several experiments mentioned

in this section. FMBZ-54 comprises of EH-TBB:BEH-TEBP in an approximately 4:1 ratio

(Bearr et al. 2012).

3.1 BTBPE

The European Food Safety Authority has published a Scientific Opinion on Emerging and

Novel Brominated Flame Retardants (BFRs) in Food, updated on 6 December 2013. It was

concluded that 1,2-bis(2,4,6-tribromophenoxy)ethane (BTBPE), with the CAS Number

37853-59-1, has a possibility to raise concerns for bioaccumulation, based on available

experimental and environmental behavior data. BTBPE was considered to be of high

persistency in the environment. In the European Union, BTBPE has been classified as low

production volume (LPV) chemical, which implies that the import or production volume is

more than 10 tonnes but less than 1000 tonnes per year. BTBPE is in pre-registration under

REACH. However, no registration dossier has been made publicly available. There is very

limited information on toxicity in humans. This section will focus on relevant information on

possible toxicity in humans, therefore, ecotoxicological information will not be discussed.

3.1.1 Hazard Identification for BTBPE

Toxicokinetic information

In a study of rats given 0.05-5% 14C-BTBPE in the diet for one day by Nomeir et al (1993),

there was a very limited amount of radioactivity eliminated in urine, of less than 1% of dose

ingested, but a high percentage of faecal excretion, of 80-100% of the dose ingested. In most

of the tissues, there were undetectable levels of radiolabeled compounds. This suggested that

BTBPE gastrointestinal absorption is poor in rats. However, in rats given a diet with 500

mg/kg bw/day 14C-BTBPE for a duration of 10 days, it was found that the adipose tissue,

kidney, skin, the thymus contained the highest concentration. Less than 0.01 % of the dose

was found in the majority of the tissues.

40

In another study by Hakk et al. (2004), rats were given a single dose of 2 mg/kg bw 14C-

BTBPE by gavage. 100% of the dose was recovered in the faeces. The same research group

also demonstrated that elimination of radioactivity by bile was less than 1%, which suggested

that the faecal elimination was primarily from unabsorbed BTBPE. Due to this low level of

absorption, tissue level of BTBPE was low. After 72 h of the single dose given, more than

0.1% of the dose was found only in the gastrointestinal tract and carcass.

3.1.2 Toxicity and Dose-Response Information on BTBPE

LD50 for BTBPE was estimated to be >10g/kg bw for rats and dogs. No compound-related

effects were observed in rats after being fed up to 10% BTBPE in the diet, at an estimated

concentration of 35 mg/kg bw/day, for 14 days. In an inhalation study, rats inhaled BTBPE at

5 or 20 mg/liter in the atmosphere for 21 days. No gross pathological changes were observed.

However, it was observed in the lungs unspecified histopathological lesions (Matthews 1984,

cited by Nomeir et al. 1993).

The reproductive and developmental toxicity of BTBPE was studied by Egloff et al. (2011). It

has been found that there are no hatching effects in chicken. In the WHO/IPCS evaluation

(2005) for genotoxicity and carcinogenicity, BTBPE was found to be not mutagenic in Ames

test and S. cerevisiae. No information was available for BTBPE on human health endpoints.

3.2 EH-TBB

In a report published in August 2015, The USEPA has evaluated a number of novel and

emerging brominated flame retardants that have been used as alternatives for phased-out

PBDEs. 2-Ethylhexyl-2,3,4,5-tetrabromobenzoate (EH-TBB) with CAS Number 183658-27-7

was one of the brominated flame retardants evaluated. EH-TBB is not found on the OECD

eChem portal. EH-TBB is not registered under REACH, and no information on production

volume in the EU is available (EFSA 2013).

USEPA has stipulated a hazard criteria used to interpret available data and assign a hazard

level. These hazard criteria, named the “Design for the Environment Alternatives Assessment

41

Criteria for Hazard Evaluation” by USEPA were finalized in 2011. When insufficient

information is available, hazard designation would be assigned conservatively by weight of

evidence (USEPA 2015). The criteria used by USEPA to assign hazard designations is

included as a table in Appendix 2.

3.2.1 Hazard Identification for EH-TBB

Toxicokinetic Information

Experimental data with FM-550 showed that it was possible for EH-TBB to be absorbed after

oral exposure from gestation and through lactation. EH-TBB was found in the tissues of

exposed dams and pups after exposure to the FM-550 (Patisaul et al. 2013). In a study by

Patusaul et al. (2013), pregnant rats were given 0, 0.1 or 1 mg/kg bw/day FM-550 in the diet

through gestation day 8 until post-natal day 21. FM550 components, including EH-TBB and

BEH-TEBP were detected in adipose, liver, and muscle tissues of dams at post-natal day 21 at

768 ng/g w.w. for high dose, and 29.6 ng/g w.w for low dose, and less than 7 ng/g w.w. in

controls. EH-TBB was also detected in pooled post-natal day 21 pup adipose tissue.

The primary metabolite of EH-TBB was found to be tetrabromobenzoic acid (CAS number

27581-13-1) by in vitro metabolism experiments with human liver microsomes, rat liver

microsomes, rat cytosol, rat intestinal microsomes, and rat serum following exposure to EH-

TBB. Phase two metabolites of tetrabromobenzoic acid was not found (Roberts et al. 2012).

Tetrabromobenzoic acid was also detected in liver tissue of dams on post-natal day 21 in the

experiment by Patusaul et al. (2013).

Hazard Identification

EH-TBB was evaluated by USEPA, according to the hazard criteria mentioned above and

included in Appendix 2, to have low acute toxicity and low genotoxicity. However, it was

evaluated to have a moderate carcinogenicity, moderate reproductive toxicity, developmental

toxicity, neurotoxicity and repeated dose toxicity. EH-TBB was considered to have a high

persistence and high tendency for bioaccumulation in the environment.

42

3.2.2 Toxicity and Dose-Response Information for EH-TBB

For Acute Mammalian Toxicity, the Acute Oral Lethality was estimated to be LD50 >5000

mg/kg bw based on several studies. EH-TBB was estimated to have uncertain potential for

carcinogenicity, based on professional judgement and analogy with closely related chemicals.

EH-TBB is considered to have a low genotoxicity by USEPA. Gene Mutation test in vitro and

Chromosomal Aberration in vitro tests yielded negative results (Chemtura 2006).

Reproductive Toxicity

EH-TBB is considered to have a moderate reproductive toxicity by USEPA. In a 2-generation

oral gavage reproductive toxicity study in rats, no reproductive effects were identified at

doses up to 165 mg/kg bw/day. This study was done with the commercial mixture FMBZ-54,

containing EH-TBB and BEH-TEBP, with the larger constituent being EH-TBB. 165 mg/kg

bw/day was the highest dose tested with the mixture FMBZ-54 and was considered as the

NOAEL. No adverse effects were observed in reproductive performance and fertility. This

NOAEL falls within the range of Moderate hazard criteria set up by USEPA (MPI Research

2008a, USEPA 2015).

Developmental Toxicity

EH-TBB is considered to have a moderate developmental toxicity by USEPA. In a 2-

generation oral gavage reproductive toxicity study in rats, given 15, 50, or 165 mg/kg-day of

FMBZ-54, it was found that pups at birth had lower body weights. Body weights were also

lower throughout lactation, in both first and second generation offspring. In the first-

generation female, premating body weight was lower. At lactation day 21, spleen weights

were decreased in first generation male pups and both male and female pups in second

generation. A NOAEL of 50 mg/kg bw/day was identified from this study based on the effects

on body weight. This NOAEL falls within the range of Moderate hazard criteria. However, it

was not very clear which component or components of the commercial mixture had caused

the observed developmental effects. LOAEL was estimated to be 165 mg/kg bw/day based on

this study. (MPI Research 2008a, USEPA 2015).

In an unpublished prenatal study by MPI Research (2008b), rats were exposed to 0, 50, 100,

300 mg/kg bw/day FMBZ-54 mixture on gestation days 6-19. There was increased incidence

of dams with sparse hair in the abdomen, lower gestation body weight, and lower food

43

consumption during gestation at doses higher than or equal to 100 mg/kg bw/day. At 100

mg/kg bw/day, lower fetal weight was observed. Incidence of fused cervical vertebral neural

arches increased in fetuses at the highest dose. At highest dose, increased incidence of fetal

ossification variations, including additional ossification centres to the cervical vertebral neural

arches, incomplete ossified skull bones (jugal, parietal, and squamosal), and unossified

sternebrae were also observed. For maternal toxicity, a NOAEL of 50 mg/kg bw/day and a

LOAEL of 100 mg/kg bw/day was estimated based on the above-described effects. For

developmental toxicity, NOAEL of 50 mg/kg bw/day and LOAEL of 100 mg/kg bw/day were

estimated based on decreased fetal weight (2008b, USEPA 2015).

Neurotoxicity

EH-TBB is conservatively designated to have a moderate neurotoxicity by USEPA. There is

very limited experimental data available and no data available on neurotoxicity screening. In a

28-day sub-chronic oral toxicity study in rats treated with FM-550 in doses 0, 160, 400, 1000

mg/kg bw/day, no neurotoxic effects were reported. The NOAEL in this study was reported to

be 1000 mg/kg bw/day, which was the highest dose tested with FM-550 (Chemtura 2006).

Repeated Dose Effects

EH-TBB is considered to have a moderate repeated dose effects, designated by USEPA based

on the two developmental and prenatal study already described above (MPI Research 2008a,

2008b).

A 28-day sub-chronic oral toxicity study was performed in rats, treated with 0, 160, 400, 1000

mg/kg bw/day. Unspecified kidney effects were reported at 1000 mg/kg bw/day. No systemic

effects were observed at 160 mg/kg bw/day, and therefore a NOEL was estimated based on

this observation. Based on kidney effects, a LOAEL of 1000 mg/kg bw/day was estimated.

NOAEL was estimated at 400 mg/kg/day (Chemtura 2006).

3.3 BEH-TEBP

In a report published in August 2015, The USEPA has evaluated a number of novel and

emerging brominated flame retardants that has been used as alternatives for phased-out

PBDEs. Bis(2-ethyl-1hexyl)tetrabromophthalate (BEH-TEBP) with CAS Number 26040-51-7

was one of the brominated flame retardants evaluated.

44

In addition, BEH-TEBP is a pre-registration substance under REACH (ECHA 2016b). It has

been classified as low production volume (LPV) chemical, which implies that the import or

production volume is more than 10 tonnes but less than 1000 tonnes per year.

3.3.1 Hazard Identification for BEH-TEBP

Toxicokinetic information

Experimental data with a commercial mixture FM-550 showed that it was possible for BEH-

TEBP to be absorbed after oral exposure. BEH-TEBP was found in the tissues of exposed

dams after exposure to the commercial mixture, however, not in the pups even though

exposure has been from gestation to lactation (Patisaul et al. 2013).

In in vitro tests, mono(2-ethylhexyl)tetrabromophthalate (CAS number 61776-60-1) was

found to the primary metabolite. In rat or human subcellular fractions, no metabolites of

BEH-TEBP was found. This metabolite was formed by purified porcine carboxylesterase at a

rate of 1.08 mol/min mg/protein. No phase two metabolite of the primary metabolite was

found (Roberts et al. 2012). BEH-TEBP has not been evaluated in humans (USEPA 2015).

BEH-TEBP has been metabolized in vitro in hepatic subcellular fractions of fathead minnow,

common carp, snapping turtle and wild-type mice (Bearr et al. 2012). There was no data

available on toxicokinetic properties of the pure BEH-TEBP compound after oral, inhalation

or dermal exposure.

Hazard Identification

This FR was evaluated by USEPA to have low acute toxicity. However, it was evaluated to

have a moderate carcinogenicity, genotoxicity, reproductive toxicity, developmental toxicity,

neurotoxicity and repeated dose toxicity.

It was stated by the REACH registration applicant that conclusion cannot be drawn for

bioaccumulation potential in mammals based on the results of study.

45

3.3.2 Toxicity and Dose-Response Information for BEH-TEBP

For acute mammalian toxicity, the acute oral lethality was estimated to have an LD50 of larger

than or equal to 2000 mg/kg in rats, based on two studies (Bradford et al. 1996, Chemtura

2006). BEH-TEBP was estimated to have uncertain potential for carcinogenicity, based on

professional judgement and analogy with closely related chemicals. BEH-TEBP is considered

to have a moderate genotoxicity. In a chromosomal aberration test with human lymphocytes,

there was a weak positive result for the test material RC9927; CASRN 26040-51-7 (Purity of

BEH-TEBP > 95%) (ACC 2004). Two other in vitro chromosomal aberration assays were

performed using a component of a commercial mixture FM-550 containing BEH-TEBP,

which yielded negative results (Chemtura 2006). In an in vivo mouse micronucleus assay, it

was found that BEH-TEBP did not cause gene mutation in bacteria or chromosomal

aberration (ACC 2004).

A study submitted by a REACH registration applicant estimated an LD50 of >5000 mg/kg

bw. 5 female and 5 male rats were administered a single oral dose of 5000 mg/kg bw of BEH-

TEBP by gavage and observed for mortality and clinical signs for 14 days. No death occurred

to any animal. Body weight gain was normal and on day 15, there was no relevant necropsy

finding (ECHA 2016b).

Reproductive Toxicity

BEH-TEBP is considered to have a moderate reproductive toxicity by USEPA. In a 2-

generation oral gavage reproductive toxicity study in rats treated with 15, 50, or 165 mg/kg-

day FMBZ-54, no reproductive effects were identified at doses up to 165 mg/kg bw/day. 165

mg/kg bw/day was the highest dose tested and was considered as the NOAEL. No adverse

effects on reproductive performance or fertility in rats were observed. This NOAEL falls

within the range of Moderate hazard criteria (MPI Research 2008a, USEPA 2015).

In a 28-day repeated dose dietary toxicity study in rats given 0, 200, 2,000, and 20,000 ppm in

diet (approx. 0, 21.1, 211, 2,110 mg/kg bw/day) of test material RC9927; CASRN 26040-51-

7 (Purity of BEH-TEBP > 95%), no adverse changes in testes or ovary weights was observed.

Gross necropsy and histopathology were performed on a full complement of male and female

reproductive organs and tissues and no adverse effects were observed. However, other

46

reproductive indicators have not been examined. A NOAEL of 2000 ppm (approx. 223.4

mg/kg bw/day) for dietary toxicity was established. 2100 mg/kg bw/day was the highest dose

tested.

Developmental Toxicity

BEH-TEBP is considered to have a moderate developmental toxicity by USEPA. In a 2-

generation oral gavage reproductive toxicity study in rats, given 15, 50, or 165 mg/kg-day of

FMBZ-54, it was found that pups at birth had lower body weights. Body weights were also

lower throughout lactation, in both first and second generation offspring. In the first-

generation female, premating body weight was lower. At lactation day 21, spleen weights

were decreased in first generation male pups and both male and female pups in second

generation. A NOAEL of 50 mg/kg bw/day was identified from this study based on the effects

on body weight. This NOAEL falls within the range of Moderate hazard criteria. However, it

was not very clear which component or components of the commercial mixture had caused

the observed developmental effects. LOAEL was estimated to be 165 mg/kg bw/day based on

this study. (MPI Research 2008a, USEPA 2015).

In an unpublished prenatal study by MPI Research (2008b), rats were exposed to 0, 50, 100,

300 mg/kg bw/day FMBZ-54 mixture on gestation days 6-19. There was increased incidence

of dams with sparse hair in the abdomen, lower gestation body weight, and lower food

consumption during gestation at doses higher than or equal to 100 mg/kg bw/day. At 100

mg/kg bw/day, lower fetal weight was observed. Incidence of fused cervical vertebral neural

arches increased in fetuses at the highest dose. At highest dose, increased incidence of fetal

ossification variations, including additional ossification centres to the cervical vertebral neural

arches, incomplete ossified skull bones (jugal, parietal, and squamosal), and unossified

sternebrae were also observed. For maternal toxicity, a NOAEL of 50 mg/kg bw/day and a

LOAEL of 100 mg/kg bw/day was estimated based on the above-described effects. For

developmental toxicity, NOAEL of 50 mg/kg bw/day and LOAEL of 100 mg/kg bw/day were

estimated based on decreased fetal weight (2008b, USEPA 2015).

In a study submitted by REACH registration applicant, pregnant rats were administered BEH-

TEBP by oral gavage at dose levels of 250, 500 or 1000 mg/kg bw/day during gestation. No

effect was found on body weight development and dietary intake. A NOEL for maternal

47

toxicity was established at 1000 mg/kg bw/day at the highest dose tested. No relevant adverse

effects were observed in the offspring. The NOEL for developmental toxicity was therefore

1000 mg/kg bw/day at the highest dose tested (ECHA 2016b).

Neurotoxicity

BEH-TEBP is conservatively designated to have a moderate neurotoxicity by USEPA. There

is very limited experimental data available and no data available on neurotoxicity screening.

In a 28-day sub-chronic oral toxicity study in rats treated with FM-550 in doses 0, 160, 400,

1000 mg/kg bw/day, no neurotoxic effects were reported. The NOAEL in this study was

reported to be 1000 mg/kg bw/day, which was the highest dose tested with FM-550

(Chemtura 2006).

Repeated Dose Effects

BEH-TEBP is considered to have a moderate repeated dose effects. In a 28-day dietary

toxicity study, a small decrease of body weight, as well as decreased phosphorus and calcium

levels in female rats was observed. A LOAEL of 2110 mg/kg bw/day was estimated. A

NOAEL was identified as 211mg/kg bw/day. A moderate hazard was designated

conservatively (ACC 2004).

In a 2-generation oral reproductive toxicity study in rats, a NOAEL of 50 mg/kg bw/day was

estimated based on reduced body weight or body weight gain during premating period in

parental F0 and F1 female rats dosed with 165 mg/kg bw/day of the same commercial

mixture. LOAEL was determined to be 165 mg/kg bw/day (Chemtura 2006).

In a repeated-dose oral toxicity study submitted by REACH registration applicant, three

groups of ten male and ten female rats received BEH-TEBP (Tradename FR-45B) by diet at

200, 2000 or 20000 ppm concentrations (= approx. 21.97, 223.4 or 2331 mg/kg bw/day) for

four weeks. A similar control group received no treatment. A positive control group with five

males and five female rats received di-2-ethyl hexyl phthalate (DEHP) by diet at

concentration 15000 ppm for four weeks. Dietary administration of BEH-TEBP to rats at the

highest dose of 20000 ppm produced only minor changes, namely, there was a slightly lower

48

overall bodyweight gain for female rats. Marginally low alanine amino-transferase activities

were also observed in females receiving the highest dose. Marginally low phosphorus

concentrations were seen in all females and males receiving the highest dose. No evidence of

toxicity was observed at 200 or 2000 ppm. BEH-TEBP did not cause any toxicity in the liver

or testes observed in positive controls given DEHP, namely peroxisome proliferation. The

NOAEL was conservatively estimated at 2000 ppm, i.e. 223.4 mg/kg bw/day for the above-

described observed effect. This study and NOAEL has been used by the REACH registrant

and ECHA to estimate the DNEL for long-term oral exposure.

49

4 EXPOSURE ASSESSMENT

4.1 Available Relevant Exposure Information

Ali et al. (2011) estimated exposure to FRs via ingestion of indoor dust by assuming 100%

absorption of intake, as it was also done in a previous study by Jones-Otazo et al. 2005. The

amount of dust ingestion was assumed to be of an average 50 mg per day for toddlers and 20

mg per day for adults. For high dust ingestion, 200 mg per day was assumed for toddlers and

50 mg per day for adults. “Low”, “Typical” and “High” exposure scenarios for each

microenvironment were estimated using 5th percentile, median, and 95th percentile

concentrations in the dust samples. Overall exposure to FR by dust ingestion was calculated

according to the estimated relative time spent in each microenvironment for toddlers and

adults.

In the Ali study, in a typical exposure scenario, exposure to BTBPE was 0.01 ng/kg bw/day

for mean dust ingestion and 0.05 ng/kg bw/day for high dust ingestion for toddlers at the

median. For non-working adult, the typical exposure was 0.02 ng/kg bw/day for mean dust

ingestion and 0.06 ng/kg bw/day for high dust ingestion at the 95th percentile (below LOQ at

median). For working adult, the typical exposure was 0.04 ng/kg bw/day for mean dust

ingestion and 0.08 ng/kg bw/day for high dust ingestion at the 95th percentile (below LOQ at

median).

Exposure to EH-TBB was 0.02 ng/kg bw/day for mean dust ingestion and 0.08 ng/kg bw/day

for high dust ingestion for toddlers at the median. For non-working adult, the typical exposure

was 0.02 ng/kg bw/day for mean dust ingestion and 0.05 ng/kg bw/day for high dust ingestion

at the 95th percentile (below LOQ at median). For working adult, the typical exposure was

also 0.02 ng/kg bw/day for mean dust ingestion and 0.05 ng/kg bw/day for high dust ingestion

at the 95th percentile (below LOQ at median).

Exposure to BEH-TEBP was 0.10 ng/kg bw/day for mean dust ingestion and 0.40 ng/kg

bw/day for high dust ingestion for toddlers at the median. For non-working adult, the typical

exposure was 0.13 ng/kg bw/day for mean dust ingestion and 0.32 ng/kg bw/day for high dust

ingestion at the 95th percentile (below LOQ at median). For working adult, the typical

50

exposure was 0.01 ng/kg bw/day for mean dust ingestion and 0.02 ng/kg bw/day for high dust

ingestion at the median.

4.2 Brominated Flame Retardants Measured in Children’s Room in Kuopio, Finland

The Institute for Health and Welfare investigated the levels of novel and emerging flame

retardants in the indoor dust of 40 children’s room in the city of Kuopio, Finland. This study

has not been published. This section presents a part of the result from this study. The levels of

brominated flame retardants ab-DBE-DBCH, PBT, TBP-DBPE, EH-TBB, BTBPE, BEH-

TEBP will be presented in this section.

Indoor dust samples were collected through 2014-2015 with nylon socks by vacuum cleaner

from the floor of children’s rooms in 40 homes in and around the city of Kuopio, with an

average area of 8-10 m2. Dust samples were sieved through standard tea filters to remove

large particles and hair. Fifty mg of dust was weighed for each sample. Samples were

analysed with method described in section 2. Table 15 Shows the level of brominated flame

retardants found in children’s rooms in Kuopio.

Table 15. Levels of BFR found in children’s rooms in Kuopio (n=40).

Compound

DBE-

DBCH PBT

TBP-

DBPE

EH-

TBB

BTBP

E

BEH-

TEBP

Mean (ng/g) 0.554 1.135 1.464 6.030 2.722 242.715

Median 0.420 0.644 0.474 3.688 1.271 106.306

STDEV 0.410 2.263 3.418 5.818 4.071 354.452

5th Percentile 0.234 0.220 0.082 0.911 0.372 22.859

95th Percentile 1.241 1.851 5.914 17.913 6.708 887.262

Table 16 shows the levels of the above six BFRs detected in recent studies in homes

(bedrooms) in Boston, USA (Stapleton et al. 2008), Belgian homes (Ali et al. 2011), homes in

New Zealand (Ali et al. 2012), home in Romania (Dirtu et al. 2012), homes in Norway

(Cequier et al. 2014), homes in Durham, USA (Stapleton et al. 2014) and homes in Canada

(Fan et al. 2016). Not all six BFRs have been analysed in every study.

From these studies, it can be observed that the levels of EH-TBB seem comparable between

the THL study and from levels measured in New Zealand and Romanian homes. The levels of

51

BTBPE seem comparable across all studies. However, all studies show a much bigger range,

in particular those measured from homes in Boston, USA, Belgium and Canada. The levels of

BEH-TEBP from the THL study is comparable to those measured in bedrooms in Boston,

USA, Belgian homes, and Norwegian homes. The level of BEH-TEBP, as well as EH-TBB

measured from homes in Durham, USA appear to be considerably higher than that from the

THL study. The levels of FR measured in Canadian homes have a relatively wide range given

a much larger sample size of n=351.

52

Table 16. Concentrations (ng/g) of BFR in indoor dust in the THL study and those reported in other studies from Europe and USA. Results

expressed as median (range).

Reference Sample

Origin

Particle

Size (µm)

Sample

Mass (g)

Year of

Sampling

No. of

Samples (n)

DBE-

DBCH

PBT TBP-

DBPE

EH-TBB BTBPE BEH-

TEBP

THL Study Kuopio,

Finland

<500 0.05 2014-2015 40 0.42

(0.202-

2.17)

0.644

(0.140-

14.5)

0.474

(0.054-

19.4)

3.69

(0.586-

22.5)

1.27

(0.270-

24.9)

106 (12-

1930)

Stapleton et

al. 2008

Boston,

MA, USA

<500 ~0.3 2006 14 90.4*

(<10.6-

378)

47.8* (1.6-

789)

105* (1.5-

763)

Ali et al.

2011

Belgium <500 ~0.075 2010 39 1 (<2 –

436)

2 (<0.5-

1019)

13 (<20-

1286)

Ali et al.

2012

New

Zealand

<500 ~0.075 2008 34 2 (<2-

2285)

2 (<2-175) 12 (<2–

640)

Dirtu et al.

2012

Iasi,

Romania

<500 ~0.075 2010 47 <2 (<2-21) 4 (<2-90) 20

Cequier et al.

2014

Oslo,

Norway

<3000 >0.1 2012 48 ∼2 ng/g ∼3 ng/g ∼5 ng/g 78.5

Stapleton et

al. 2014

Durham,

NC, USA

<500 ~0.1 2009-2010 30 97 (6.0-

2430)

604 (82.9-

20960)

Fan et al.

2016

Canada <80 ~0.1 2007-2010 351 <0.6 (<0.6-

46)

Not

Detected

104 (<1.5-

13000)

8.5 (<1.7-

2390)

* Geometric mean

53

4.3 Exposure Estimation

This exposure estimation, shown below in Table 17, was based on the study by Ali et al.

(2011). Levels of BFR in indoor dust have been taken from the unpublished THL study

presented in section 4.2. In this exposure estimation, 100% absorption of intake is assumed, as

is done by Ali et al. (2011) and Jones-Otazo et al. (2005). In both aforementioned studies,

adult dust ingestion was assumed to be 20 mg/day for average ingestion, and 50 mg/day for

high ingestion. Toddler dust ingestion was assumed to be 50 mg/day for average ingestion,

and 200 mg/day for high ingestion.

Adult body weight was assumed to be 70kg, and toddler body weight assumed to weigh 10kg,

based on dose exposure calculation guidelines developed by the United States

Agency for Toxic Substances and Disease Registry (ATSDR 2005).

Table 18 presents a comparison of exposure values estimated from THL studies to studies by

Ali et al. (2011) estimated from Belgian homes & Ali et al. (2012) estimated from homes in

New Zealand. The exposure to EH-TBB and BTBPE are comparable across the three studies.

Toddler exposure to all three BFRs at 95th percentile in Belgian homes tend to be higher

compared to the other two studies. It is observed that exposure to BEH-TEBP from the THL

study, with samples taken from children’s rooms in Kuopio, is observed to be at least 2 times

higher than the other two studies for both toddler and adult exposure for all exposure ranges.

This may be due to a high median of 106 ng/g BEH-TEBP indoor dust level from the THL

study used for the exposure estimation. The median used for the Ali et al. studies were 13

ng/g for Belgian homes (2011) and 12 ng/g for New Zealand homes (2012). The range of

BEH-TEBP levels from the Ali et al. (2011) study of Belgian homes is comparable to that of

the THL study. The range from the 2012 study of New Zealand homes is smaller, as can be

seen in Table 16 above.

54

Table 17. Estimated oral exposure to BFRs through dust ingestion.

Toddler (10 kg bw) Adult (70 kg bw)

(ng/kg bw/day) 5th Percentile 95th Percentile Median 5th Percentile 95th Percentile Median

DBE-DBCH

Mean dust ingestion 0.001 0.006 0.002 0.000 0.000 0.000

High dust ingestion 0.005 0.025 0.008 0.000 0.001 0.000

PBT



TBP-DBPE



EH-TBB



BTBPE



BEH-TEBP



55

Table 18. Comparison of exposure values estimated from THL studies to studies by Ali et al. (2011 & 2012).

Toddler (10 kg bw) Adult (70 kg bw)

(ng/kg bw/day) 5th Percentile 95th Percentile Median 5th Percentile 95th Percentile Median

EH-TBB (Ali et al. 2011)



EH-TBB (Ali et al. 2012)

Mean dust ingestion 0.010 0.050 0.010 <0.01 <0.01 <0.01

High dust ingestion 0.020 0.200 0.040 <0.01 0.010 <0.01

EH-TBB (THL)



BTBPE (Ali et al. 2011)



BTBPE (Ali et al. 2012)

Mean dust ingestion <0.01 0.050 <0.01 <0.01 <0.01 <0.01

High dust ingestion 0.010 0.200 0.010 <0.01 0.010 <0.01

BTBPE (THL)



BEH-TEBP (Ali et al. 2011)



BEH-TEBP (Ali et al. 2012)

Mean dust ingestion <0.01 0.260 0.050 <0.01 <0.01 0.020

56

High dust ingestion 0.100 1.020 0.190 <0.01 0.040 0.010

BEH-TEBP (THL)



57

5 RISK CHARACTERISATION FOR BEH-TEBP

Based on the exposure estimation from Table 16 for BEH-TEBP, a Risk Characterisation

Ratio has been calculated based on the derived no-effect level (DNEL) of 0.37 mg/kg bw/day

(ECHA 2016b) for oral exposure in the general population. This DNEL has been derived for

long-term oral exposure according to a repeated dose oral toxicity study described in Section

3.3.2. The RCR calculated are shown in Table 18 below.

Table 18. RCR Calculation for BEH-TEBP based on DNEL of 0.37 mg/kg bw/day for oral

exposure in the general population. Toddler (10 kg bw) Adult (70 kg bw)

5th Percentile 95th Percentile Median 5th Percentile 95th Percentile Median

BEH-TEBP



6. DISCUSSION

Summary and Data Gaps for BTBPE

BTBPE is primarily eliminated in the faeces. It is shown that BTBPE has poor gastrointestinal

absorption in rats, however, a small percentage (<0.01%) of BTBPE may persist in the

adipose tissue, kidney, skin, and thymus. BTBPE has a high LD50 of >10g/kg bw. BTBPE

was found not mutagenic in Ames test and in yeast. There are also no hatching effects in

chicken. Based on available information, unspecified histopathological lesions in the lungs

has been observed in rats after inhalation exposure for 21 days. Based on this limited

information, inhalation exposure in humans may be important in terms of possible health risk.

However, there is no information available for BTBPE on any human health endpoints.

For BTBPE, there is very limited published information regarding human health risk and

human health end-points. Therefore, information on carcinogenicity, reproductive toxicity,

developmental toxicity, neurotoxicity and repeated-dose toxicity are not available.

Exposure to BTBPE is among the lowest in the list of BFR considered, however, toddler

exposure to BTBPE for high dust ingestion in the 95th percentile can exceed 1 ng/kg bw/day.

58

Even though BTBPE is considered a LPV chemical, more information on human health-

related end-points should be made available.

Summary and Data Gaps for EH-TBB

Toxicokinetic study was conducted with the commercial mixture FM-550 containing a

mixture of EH-TBB and BEH-TEBP. Absorption was possible through oral exposure from

gestation and through lactation in rats. EH-TBB and BEH-TEBP were detected in adipose,

liver and muscle tissues of dams, and in pup adipose tissue on post-natal day 21. The primary

metabolite of EH-TBB was found in the liver tissue of dams on post-natal day 21. EH-TBB

has an LD50 of >5000 mg/kg bw. The commercial mixture FMBZ-54 (a mixture of EH-TBB

and BEH-TEBP) was found to have no adverse effect in reproductive performance and

fertility at 165 mg/kg bw/day. However, in a prenatal study with rats, it was found that there

was higher incidence of dams with sparse hair in the abdomen, lower gestation body weight,

and lower food consumption during gestation at dose 100 mg/kg bw/day. At this dose level,

lowered fetal weight was observed. At 300 mg/kg bw/day, there was increased incidence of

fused cervical vertebral neural arches in fetus, and other developmental effects. With the

consideration of human health, it is significant that absorption of EH-TBB was found to be

possible through lactation. Deposition of EH-TBB was also possible in adipose, liver and

muscle tissues. Moreover, at high dose, adverse effects were observed in pregnant rat dams

and in rat foetal development. Therefore, foetal exposure and neonatal exposure through

lactation, as well as accumulation in adipose can be a human health concern.

There was only one study reported for neurotoxicity with no effect observed at 1000 mg/kg

bw/day. Apart from this, there is very limited experimental data available and no data

available on neurotoxicity screening. Details on kidney effects from the 28-day sub-chronic

oral toxicity study performed by Chemtura (2006) has not be made publicly available.

Exposure to EH-TBB is among the lowest in the list of BFR considered, however, toddler

exposure to EH-TBB for high dust ingestion in the 95th percentile can exceed 1 ng/kg

bw/day. Given the above possible human health concern, especially in feotal development,

neurotoxicity studies would be informative. Human biomonitoring of breast milk would also

be relevant.

59

Summary and Data Gaps for BEH-TEBP

Toxicokinetic study was conducted with the commercial mixture FM-550. BEH-TEBP was

found in the tissues of exposed dams after exposure to the commercial mixture, however, not

in the pups even though exposure has been from gestation to lactation. BEH-TEBP has an

LD50 of >5000 mg/kg bw. The REACH registrant for BEH-TEBP has provided studies

conducted with the compound BEH-TEBP of >95% purity. This test material was found to

cause no adverse reproductive effects in rats, with an estimated NOAEL of approximately

223.4 mg/kg bw/day dietary exposure. No developmental effects were observed at 1000

mg/kg bw/day, which was the highest dose tested and the NOEL. For repeated-dose oral

toxicity, NOAEL was conservatively estimated at 2000 ppm, i.e. 223.4 mg/kg bw/day for

minor observed effects. There was a slightly lower overall bodyweight gain for female rats.

Marginally low alanine amino-transferase activities were also observed in females receiving

the highest dose of 2331 mg/kg bw/day Marginally low phosphorus concentrations were seen

in all females and males receiving the highest dose. This study and the estimated NOAEL has

been used by the REACH registrant and ECHA to estimate the DNEL for long-term oral

exposure.

For BEH-TEBP, there was only one study reported for neurotoxicity with no effect observed

at 1000 mg/kg bw/day. Apart from this, there is very limited experimental data available and

no data available on neurotoxicity screening. Neurotoxicity study has not been included in the

registration dossier.

Among international results as well as results from THL (Table 16), BEH-TEBP was present

at the highest level among the six BFR measured. The estimated exposure was also highest

(Table 17, Table 18). Based on the THL study, the estimated exposure at median

concentration for mean dust ingestion was 0.532 ng/kg bw/day for toddlers and 0.030 ng/kg

bw/day for adults. However, for high dust ingestion at 95th percentile concentration, the

exposure can reach 17.7 ng/kg bw/day for toddlers and 0.634 ng/kg bw/day for adults. BEH-

TEBP should be of particular concern due to the relatively high level of exposure. Therefore,

information on neurotoxicity would be relevant. Human biomonitoring of serum and breast

milk concentration of BEH-TEBP would also be relevant.

60

Discussion on Risk Characterisation

Based on the DNEL of 0.37 mg/kg bw/day for oral exposure in the general population, the

RCR calculated for all exposure scenarios for both toddlers and adults are well below 1,

indicating that the risk from oral exposure through house dust is adequately controlled.

However, it must be kept in mind that the sample size of 40 from children’s rooms in Kuopio,

Finland is insufficient to represent the Finnish population, nor the population of the city of

Kuopio. Moreover, RCR has been calculated based on the exposure estimation through house

dust alone, combined exposure has not been considered.

The DNEL employed in the risk characterization is considered reliable. Toxicological

information from the REACH registration dossier for BEH-TEBP are based on study with

high reliability, with a denomination of 1 – reliable without restrictions, or 2 – reliable with

restrictions.

Further Considerations

Many of the toxicological studies have been conducted with commercial mixtures of EH-TBB

and BEH-TEBP, namely FM-550 and FMBZ-54. In the REACH registration dossier for BEH-

TEBP, tests have been conducted with the compound BEH-TEBP of >95% purity. This test

material was found to cause no adverse reproductive effects in rats at 223.4 mg/kg bw/day

dietary exposure for four weeks, and no developmental effects were observed at 1000 mg/kg

bw/day when exposed during gestation. However, the commercial mixture FMBZ-54 was

found to cause maternal toxicity at 100 mg/kg bw/day and foetal developmental effects at

300 mg/kg bw/day when exposed during gestation day 6-19.

It is reasonable to assume that since these commercial mixtures are being applied to furniture,

humans would be exposed to flame retardants as mixtures as well. Moreover, in many studies

conducted with commercial mixtures, it is unclear which components of the mixture may be

driving the observed adverse effects (USEPA 2015). Therefore, it is important to consider

human exposure to flame retardants as mixtures, and the possibility of synergistic effect of

these chemical mixtures. Further studies in this respect would be very informative.

61

7. SUMMARY AND CONCLUSIONS

Flame retardants (FR) have been added to a wide range of industrial and commercial products

as a result of flammability standards, therefore FRs are ubiquitous in the indoor environment.

FRs can accumulate in indoor dust, and levels of FRs present in house dust can be an

indication of human exposure to FRs. Brominated flame retardants (BFR) and

Organophosphorous Flame Retardants (OPFR) have been developed to replace banned

Polybrominated diphenyl ethers (PBDE). However, the toxicity of these novel and emerging

FRs to human health, and possible tendency for bio-accumulation have become a concern.

Method development and validation for analysis of flame retardants in house dust that was

performed between May-August 2015 based on the method developed by Van den Eede et al.

2012. The principle aim of this method development is to develop and validate an effective

analysis method for indoor dust containing selected novel and emerging BFR, OPFR, as well

as PBDEs. In the second part of this thesis, a human health risk assessment for one of the

above selected BFRs, namely BEH-TEBP was performed based on currently available

scientific literature and estimated exposure from dust ingestion.

There was insufficient toxicological and exposure information available for ab-DBE-DBCH,

PBT, TBP-DBPE to perform a hazard identification. Relevant information available for the

substances BTBPE, EH-TBB and BEH-TEBP were summarized in Section 3. An exposure

estimation has been performed based on the amount of BFR detected from a recent

unpublished study by THL. Based on this exposure estimation, Risk Characterisation Ratio

(RCR) has been calculated for BEH-TEBP according to the derived no-effect level (DNEL)

for oral exposure.

Dichloromethane was chosen to be the most suitable extraction solvent due to ease of

preparation and use. Direct pouring of 4ml Dcm extract to cleanup columns was chosen for

future implementation. It was also concluded that 100% Dcm was the most suitable solvent

for the elution of Fraction 1 from the dual clean-up columns with Florisil column on top of

H2SO4-silica column was used for Fraction 1 only. With this solvent most of the BFRs could

be separated from the OPFRs. 10% Acetone-Dcm was determined to be the most suitable

solvent for the elution of Fraction 2, where all of OPFR, could be eluted, except that 50% of

62

TBOEP still remained in the column. BEH-TEBP was eluted in Fraction 2 together with

OPFR. Settled dust was considered to be a preferable matrix.



values, or values from calibration standards. For the OPFRs, chromatography peak sizes

tended to be too small in the calibration standard for all compounds, and for some compounds

in the Internal Standard solution added to samples.

For the validation, Limit of Quantification (LOQ) & Measurement Uncertainties were

acceptable for all BFRs. LOQ of BFRs ranged between 0.5 – 5.0 ng/g. For OPFR, LOQ

ranged between 6.9-613 ng/g. Precision for OPFR was good, but accuracy was poor compared

to other labs’ analysis result for the certified material for indoor dust SRM 2585. Therefore,


this thesis. Exposure estimation was based on the study by Ali et al. (2011), where 100%

absorption of intake is assumed. Adult dust ingestion was assumed to be 20 mg/day for

average ingestion, and 50 mg/day for high ingestion. Toddler dust ingestion was assumed to

be 50 mg/day for average ingestion, and 200 mg/day for high ingestion.

For the purpose of estimating exposure, unpublished results from the National Institute for

Health and Welfare (THL) were used. The detected amount of BEH-TEBP from children’s

room in Kuopio, Finland (n=40) had a median of 106.3 ng/g, with a range of 22.8 ng/g –

887.2 ng/g (5th to 95th percentile). A Risk Charactrisation Ratio has been calculated based on

the DNEL of 0.37 mg/kg bw/day (ECHA 2016b) for oral exposure in the general population.

All RCR derived for given exposure scenarios are less than 1, meaning that the risk is

adequately controlled. However, it must be kept in mind that combined exposure has not been

taken into consideration.

63

8. REFERENCES

ACC. 2004. High Production Volume (HPV) Challenge Program. Test plan for phthalic acid

tetrabromo bis 2-ethylhexyl ester (CAS# 26040-51- 7). Robust summaries & test plans:

Diisopropyl ether. American Chemistry Council. Submitted under the HPV Challenge

Program. Summary available at:

<http://www.epa.gov/HPV/pubs/summaries/phthacid/c15484tp.pdf>.

Ali N, Harrad S, Goosey E, Neels H and Covaci A. 2011. “Novel” brominated flame

retardants in Belgian and UK indoor dust: Implications for human exposure. Chemosphere

83: 1360-1365.

Ali N, Dirtu AC, Van den Eede N, Goosey E, Harrad S, Neels H, Mannetje t A, J C, Douwes

J, Covaci A. 2012a. Occurrence of alternative flame retardants in indoor dust from New

Zealand: indoor sources and human exposure assessment. Chemosphere 88: 1276–82.

Bearr JS, Mitchelmore CL, Roberts SC, et al. 2012. Species specific differences in the in vitro

metabolism of the flame retardant mixture, Firemaster(R) BZ-54. Aquatic Toxicology 124-

125:41-47.

Bradford L, Pinzoni E, Wuestenenk J. 1996. The Effect of Fogging of Common FR Additives

in Flexible Foam. Proceedings of the Polyurethane Foam Association, October 17 & 18,

1996. Akzo Nobel Central Research.

Bureau Veritas Consumer Product Service, USA. 2014. California Enacts Upholstered

Furniture Flame Retardant Labeling Law. Available at: <http://www.bureauveritas.com/>.

Accessed 10 May 2015.

Center for Environmental Health (CEH), USA. 2015. TB117-2013 State of California FAQ.

Available at: <http://www.ceh.org/campaigns/flame-retardants/faqs/>. Accessed 8 April

2015.

Cequier E, Ionas AC, Covaci A, Marce RM, Becher G. and Thomsen C. 2014. Occurrence of

a broad range of legacy and emerging flame retardants in indoor environments in Norway.

Environmental science & technology, 48(12): 6827-6835.

Chemtura. 2006. Material Safety Data Sheet FIREMASTER 550. Chemtura Product Safety

Group.

Chemtura. 2008. Material Safety Data Sheet FIREMASTER BZ-54 HP.

Chemtura. 2013. Material Safety Data Sheet for Firemaster BZ-54. Chemtura Corporation.

Chen A et al. 2014. Prenatal Polybrominated Diphenyl Ether Exposures and Development in

U.S. Children through 5 Years of Age: The HOME Study.

Environmental Health Perspectives 122(8): 856-862.

64

Chen SJ, Ma YJ, Wang J, Chen D, Luo XJ and Mai BX. 2009. Brominated Flame Retardants

in Children’s Toys: Concentration, Composition, and Children’s Exposure and Risk

Assessment. Environmental Science and Technology 43: 4200-4206.

Dirtu AC, van den Eede N, Ali N, Neels H and Covaci A. 2011. Profile for chlorinated and

brominated organic contaminants in indoor dust. case study for Iasi, Eastern Romania.

Organohalogen Compounds 73: 3-12.

Egloff C, Crump D, Chiu S, Manning G, McLaren KK, Cassone CG, Letcher RJ, Gauthier LT

and Kennedy SW. 2011. In vitro and in ovo effects of four brominated flame retardants on

toxicity and hepatic mRNA expression in chicken embryos. Toxicology Letters 207: 25-33.

European Chemicals Agency (ECHA). 2016a. Guidance on Information Requirements

and Chemical Safety Assessment Part E: Risk Characterisation. Available from: <

https://echa.europa.eu/documents/10162/13632/information_requirements_part_e_en.pdf>.

Accessed: 2 April 2017.

European Chemicals Agency (ECHA). 2016b. Registration Dossier: Bis(2-ethylhexyl)

tetrabromophthalate. Available from: < https://echa.europa.eu/registration-dossier/-

/registered-dossier/10235/1>. Accessed: 2 April 2017.

European Flame Retardants Association (EFRA). 2015. Fire Protection. Available at

<http://www.cefic-efra.com/index.php/en/fire-protection>. Accessed 8 April 2015.

European Food Safety Authority. 2012. Scientific Opinion on Emerging and Novel

Brominated Flame Retardants (BFRs) in Food. EFSA Journal 2012;10(10):2908. [133 pp.]

doi:10.2903/j.efsa.2012.2908. Available from:< http://www.efsa.europa.eu/efsajournal>.

Accessed: 1 March 2017.

European Food Safety Authority (EFSA). 2016. EU Framework of Brominated Flame

Retardants. Available at < http://www.efsa.europa.eu/en/topics/topic/bfr>. Accessed 11

October 2016.

European Union (EU). 2011. Restriction of Hazardous Substances Directive 2011/65/EU. The

European Parliament and the Council of the European Union, Strasbourg.

Fan X, Kubwabo C, Rasmussen PE and Wu F. 2016. Non-PBDE halogenated flame

retardants in Canadian indoor house dust: sampling, analysis, and occurrence. Environmental

Science and Pollution Research 23: 7998-8007.

Hakk H, Larsen G and Bowers J. 2004. Metabolism, tissue disposition, and excretion of 1,2-

bis(2,4,6- tribromophenoxy)ethane (BTBPE) in male Sprague-Dawley rats. Chemosphere 54:

1367-1374.

Herbstman JB et al. 2010. Prenatal Exposure to PBDEs and Development. Environmental

Health Perspectives 118(5): 712-719.

Jiang H et al. 2014. Daily intake of polybrominated diphenyl ethers via dust and diet from an

e-waste recycling area in China. Journal of Hazardous Materials 276(2014): 35-42.

http://www.efsa.europa.eu/efsajournal

65

Little JC, Weschler CJ, Nazaroff WW, Liu Z, and Cohen Hubal EA. 2012. Rapid methods to

estimate potential exposure to semivolatile organic compounds in the indoor environment.

Environmental Science & Technology 46:11171–78. doi:10.1021/es301088a.

MPI Research. 2008a. CN-2065: An oral two-generation reproduction and fertility study in

rats. MPI Research Inc.

MPI Research. 2008b. CN-2065: Prenatal developmental toxicity study in rats. MPI Research

Inc.

Munn SJ. 2007. The REACH Tool: Derived no effects level (DNEL). BAuA Conference on

Occupational Limit Values for Hazardous substances Dortmund, Germany, 7-8th May 2007.

Available from:

<https://www.baua.de/DE/Angebote/Veranstaltungen/Dokumentationen/Gefahrstoffe/pdf/Gre

nzwert-Tagung-2007-03.pdf?__blob=publicationFile>. Accessed: 13 April 2017.

Ni K et al. 2013. A review of human exposure to polybrominated diphenyl ethers (PBDEs) in

China. International Journal of Hygiene and Environmental Health 216(2013): 607-623.

Nomeir AA, Markham PM, Ghanayem BI and Chadwick M. 1993. Disposition of the flame-

retardant 1,2-bis(2,4,6-tribromophenoxy)ethane in rats following administration in the diet.

Drug Metabolism and Disposition, 21, 209-214.

Norrgran, J et al. 2015. Higher PBDE Serum Concentrations May be Associated with Feline

Hyperhyroidism in Swedish Cats. Environmental Science & technology 49(8): 5104-5114).

Patisaul HB, Roberts SC, Mabrey N, et al. (2013) Accumulation and endocrine disrupting

effects of the flame retardant mixture Firemaster 550 in rats: an exploratory assessment. J

Biochem Mol Toxicol 27(2):124-36.

Roberts SC, Macaulay LJ, Stapleton HM (2012) In vitro metabolism of the brominated flame

retardants 2-ethylhexyl-2,3,4,5-tetrabromobenzoate (TBB) and bis(2-ethylhexyl) 2,3,4,5-

tetrabromophthalate (TBPH) in human and rat tissues. Chem Res Toxicol 25(7):1435-1441.

Société Générale de Surveillance (SGS). 2017. EU Regulates Deca-BDC Under REACH.

Available at: < http://www.sgs.com/en/news/2017/02/safeguards-02717-eu-regulates-deca-

bde-under-reach>. Accessed 12 April 2017.

Stapleton HM, Allen JG, Kelly SM, Konstantinov A, Klosterhaus S, Watkins D, McClean

MD, Webster TF. 2008. Alternate and new brominated flame retardants detected in US house

dust. Environ. Sci. Technol. 42: 6910–6916.

SP Technical Research Institute of Sweden. 2016. Information about EN 1021-1:2014 and EN

1021-2:2014. Available at

<http://www.sp.se/en/index/services/firetest_furniture/ss_en_1021/sidor/default.aspx#sthash.8

Q1XPNDS.dpuf>. Accessed 16 June 2016.

https://www.baua.de/DE/Angebote/Veranstaltungen/Dokumentationen/Gefahrstoffe/pdf/Grenzwert-Tagung-2007-03.pdf?__blob=publicationFile

https://www.baua.de/DE/Angebote/Veranstaltungen/Dokumentationen/Gefahrstoffe/pdf/Grenzwert-Tagung-2007-03.pdf?__blob=publicationFile

http://www.sgs.com/en/news/2017/02/safeguards-02717-eu-regulates-deca-bde-under-reach

http://www.sgs.com/en/news/2017/02/safeguards-02717-eu-regulates-deca-bde-under-reach

66

United Nations (UN). 2015. Revised draft guidance for the inventory of polybrominated

diphenyl ethers under the Stockholm Convention. From the Proceedings of the Seventh

Meeting of the Conference of the Parties to the Stockholm Convention on Persistent Organic

Pollutants. Geneva, 4–15 May 2015. Available at<

http://chm.pops.int/Implementation/NIPs/Guidance/GuidancefortheinventoryofPBDEs/tabid/3

171/Default.aspx>. Accessed 11 October 2016.

United States Agency for Toxic Substances and Disease Registry (ATSDR). 2005. Public

Health Assessment Guidance Manual (2005 Update): Appendix G: Calculating Exposure

Doses. Available from:< https://www.atsdr.cdc.gov/hac/phamanual/appg.html>. Accessed: 2

April 2017.

United States Environmental Protection Agency (USEPA). 2017. Human Health Risk

Assessment. Available from:<https://www.epa.gov/risk/human-health-risk-assessment>.

Accessed: 13 April 2017.

United States Environmental Protection Agency (USEPA). 2015. Flame Retardants Used in

Flexible Polyurethane Foam: An Alternatives Assessment Update. EPA Publication 744-R-

15-002. Available from:< https://www.epa.gov/saferchoice/flame-retardants-used-flexible-

polyurethane-foam>. Accessed: 4 March 2017.

Van den Eede N et al. 2012. Multi-residue method for the determination of brominated and

organophosphate Flame retardants in indoor dust. Talanta 89(2012): 292-300.

Wang J et al. 2010. Brominated flame retardants in house dust from e-waste recycling and

urban areas in South China: Implications on human exposure. Environment International 3

(2010): 535-541.

WHO/IPCS (World Health Organization), 2005. Concise International Chemical Assessment

Document 66. 2.4.6-Tribromophenol and other simple brominated phenols. Geneva,

Switzerland. Available from:

<http://www.who.int/ipcs/publications/cicad/cicad_66_web_version.pdf>. Accessed: 2 April

2017

https://www.atsdr.cdc.gov/hac/phamanual/appg.html

https://www.epa.gov/risk/human-health-risk-assessment

https://www.epa.gov/saferchoice/flame-retardants-used-flexible-polyurethane-foam

https://www.epa.gov/saferchoice/flame-retardants-used-flexible-polyurethane-foam

http://www.who.int/ipcs/publications/cicad/cicad_66_web_version.pdf

67

APPENDICES

APPENDIX 1: Reference values for BFR and OPFR analysis in house dust

Compoun

d (Abbr.)

CAS-

Numbe

r

Full Chemical name Certifie

d Value

(SRM-2585)

ng/g

Ali 2011

(ng/g) -

Indicative Value

Van den

Eede 2011

(ng/g) - Indicative

Value

Van den

Eede 2012

(ng/g) - Measured

Value

Sahlströ

m 2012

(ng/g)

Cristale

2013

(ng/g)

Stapleto

n 2008

(ng/g)

Hoffma

n 2015

(ng/g)

Bergh 2012

(ng/g)

Fan

2016

(ng/g)

Iona

s

2013 (ng/g

)

Fromm

e 2014

(ng/g)

Ali 2012

(ng/g)

TCEP 115-

96-8

Tris(2-chloroethyl)

phosphate

700(170) 680(60) 840(60) 700 788(21)

TCIPP 13674-

84-5

Tris(1-chloropropan-2-yl)

phosphate

820(100) 860(70) 880(140) 1000 840(19)

TDCIPP 13674-

87-8

tris(1,3-

dichloroisopropyl) phosphate

2020(260) 3180(70) 1820(9

0)

2300(280) 2200 1936(95)

TDBPP 126-

72-7

Tris(2,3-dibromopropyl)

phosphate

TIBP 126-

71-6

Tri(isobutyl)phosphate <(MDL=29

0)

300 1565(388)

TNBP 126-

73-8

Tri-n-butylphosphate 180(20) 190(10) 190(20) 400 169(20)

TBOEP 78-51-

3

Tris(2-

butoxyethyl)phosphate

49,000(960

0)

63,000(200

0)

82000(6500

), 73000

71000 40,107(32

4)

EHDPP 1241-

94-7

2-Ethylhexyl diphenyl

phosphate

1300(120),

1000

1900

TEHP 78-42-

2

Tris(2-

ethylhexyl)phosphate

370(40),

330

500

TPHP 115-

86-6

Triphenyl phosphate 990(70) 1160(140) 520

(34)

1100(100) 700 1,058(85)

TMPP 1330-

78-5

Tris (methylphenyl)

phosphate

1070(110) 1140(30) 740(110)

ab-DBE-DBCH

3322-93-8

alpha/beta-Tetrabromoethylcyclohex

ane

PBT 87-83-

2

Pentabromotoluene

PBEB 7.7(0.6)

HBB 2.8(0.4) 2(0.8

)

TBP-

DBPE

35109-

60-5

2,3-Dibromopropyl 2,4,6-

tribromophenyl ether

68

EH-TBB 18365

8-27-7

2-Ethylhexyl-2,3,4,5-

tetrabromobenzoate

40 26(2) 36(2.4) 35(6) 38.8(4.

8)

40(2

)

BTBPE 37853-59-1

1,2-Bis(2,4,6-tribromophenoxy)ethane

32 39(14) 39(4.9) 76(4) 37.8(5.9)

35(4)

BEH-

TEBP

26040-

51-7

Bis(2-ethyl-

1hexyl)tetrabromophthala

te

652 574(49) 1300(94

)

857(73) 145(16.

7)

DBDPE 84852-53-9

Decabromodiphenylethane

< 20 <7.1

BDE-28 2,2',4-Tribromodiphenyl

Ether

46.9

(4.4)

32.8(1.1) 42(1)

BDE-47 2,2',4,4'-Tetrabromodiphenyl

Ether

497 (46)

409(11) 486(10)

BDE-99 2,2',4,4',5-

Pentabromodiphenyl Ether

892

(53)

742(23) 803(45)

BDE-100 2,2',4,4',6-

Pentabromodiphenyl

Ether

145

(11)

116(3) 147(5)

BDE-153 2,2',4,4',5,5'-

Hexabromodiphenyl

Ether

119 (1) 97(2) 118(8)

BDE-154 2,2',4,4',5,6'-

Hexabromodiphenyl Ether

83.5

(2.0)

77.2(2.7) 77(5)

BDE-183 2,2',3,4,4',5',6-

Heptabromodiphenyl

Ether

43.0

(3.5)

32.3(4.8) 44(4)

BDE-209 Decabromodiphenyl Ether 2510

(190)

2150(231) 2971(33

3)

* (ng/g) = (µg/kg) [a] Ali, N. et al. 2011. ‘Novel’’ brominated flame retardants in Belgian and UK indoor dust: Implications for human exposure. Chemosphere 83 (2011) 1360-1365.

[b] Cristale,J.; Lacorte,S. 2013. Development and validation of a multiresidue method for the analysis of polybrominated diphenyl ethers, new brominated and organophosphorus flame retardants in sediment, sludge and

dust. J. Chromatogr. A. 1305 (2013) 267-275.

[c] Sahlström et al. 2012. Clean-up method for determination of established and emerging brominated flame retardants in dust. Anal. Bioana. Chem. 404 (2912) 459.

[d] Stapleton H.M. et al. 2008. Alternate and new brominated flame retardants detected in U.S. house dust. Enviro. Sci. Technol. 42 (2008) 6910. [e] Van den Eede, N. et al. 2011. Analytical developments and preliminary assessment of human exposure to organophosphate flame retardants from indoor dust. Environment International 37(2) 454-461.

[f] Van den Eede, N. et al. 2012. Multi-residue method for the determination of brominated and organophosphate flame retardants in indoor dust. Talanta 89 (2012) 292-300.

[g] Hoffman, K. et al. 2015. Monitoring Indoor Exposure to Organophosphate Flame Retardants: Hand Wipes and House Dust. Env. Health. Pespec. 123(2) 160-165.

[h] Bergh, C. et al. 2012. Organophosphate and phthalate esters in standard reference material 2585 organic contaminants in house dust. Anal. Bioanal. Chem. 402 (2012) 51-59. [i] Fan, X. et al. 2016. Non-PBDE halogenated flame retardants in Canadian indoor house dust: sampling, analysis, and occurrence. Environ Sci Pollut Res (In press).

[j] Ionas A.C.; Covaci A. 2013. Simplifying multi-residue analysis of flame retardants in indoor dust. Int J Environ Anal Chem 93:1074–1083.

[k] Ali N. et al. 2012. Assessment of human exposure to indoor organic contaminants via dust ingestion in Pakistan Indoor Air 22(3) 200-211.

69

APPENDIX 2: Criteria Used by USEPA to Assign Hazard Designation

Endpoint Very High High Moderate Low Very Low

Human Health Effects

Acute mammalian toxicity

Oral median lethal

dose (LD50) (mg/kg) ≤50

>50–300 >300–2,000

>2,000

–

Dermal LD50 (mg/kg) ≤200 >200–1,000 >1,000–2,000 >2,000 –

Inhalation median

lethal concentration

(LC50) - vapor/gas

(mg/L)

≤2 >2–10 >10–20 >20 –

Inhalation LC50 -

dust/mist/ fume

(mg/L)

≤0.5 >0.5–1.0

>1–5

>5

–

70

Carcinogenicity

Carcinogenicity

Known or presumed

human carcinogen

(equivalent to Globally Harmonized

System of

Classification and

Labeling of Chemicals (GHS)

Categories 1A and

1B)

Suspected human

carcinogen

(equivalent to GHS

Category 2)

Limited or marginal

evidence of

carcinogenicity in

animals

(and inadequate

evidence in humans)

Negative studies or

robust mechanism-

based Structure

Activity Relationship

(SAR)

(as described above)

–

71

Mutagenicity/Genotoxicity

Germ cell

mutagenicity

GHS Category 1A or

1B: Substances

known to induce

heritable mutations or

to be regarded as if

they induce heritable mutations in the germ

cells of humans

GHS Category 2:

Substances which cause concern for

humans owing to the

possibility that they

may induce heritable mutations in the germ

cells of humans

OR

Evidence of mutagenicity

supported by positive

results in in vitro OR

in vivo somatic cells

of humans or animals

Negative for

chromosomal

aberrations and gene

mutations, or no

structural alerts.

--

Mutagenicity and

genotoxicity in

somatic cells

Evidence of mutagenicity

supported by positive

results in in vitro

AND in vivo somatic

cells and/or germ

cells of humans or

animals

72

Reproductive toxicity

Oral (mg/kg/day) –

<50 50–250

>250-1,000

>1,000

Dermal (mg/kg/day) – <100 100–500 >500-2,000 >2,000

Inhalation - vapor,

gas (mg/L/day) – <1 1–2.5 >2.5-20 >20

Inhalation -

dust/mist/fume

(mg/L/day)

– <0.1 0.1–0.5 >0.5-5 >5

Developmental toxicity

Oral (mg/kg/day) – <50 50–250 >250-1,000 >1,000

Dermal (mg/kg/day) – <100 100–500 >500-2,000 >2,000

Inhalation - vapor,

gas (mg/L/day) – <1 1–2.5 >2.5-20 >20

Inhalation -

dust/mist/fume

(mg/L/day)

–

<0.1 0.1–0.5 >0.5-5 >5

73

Neurotoxicity


<10 10–100

>100

–

Dermal (mg/kg/day) – <20 20–200 >200 –

Inhalation - vapor,

gas (mg/L/day) - <0.2 0.2–1.0 >1.0 -

Inhalation -

dust/mist/fume

(mg/L/day)

- <0.02 0.02–0.2 >0.2 -

74

Repeated-dose toxicity


<10 10–100

>100

–

Dermal (mg/kg/day) – <20 20–200 >200 –

Inhalation - vapor, gas

(mg/L/day) - <0.2 0.2–1.0 >1.0 -

Inhalation - dust/mist/fume

(mg/L/day) - <0.02 0.02–0.2 >0.2 -

*Very High or Very Low designations (if an option for a given endpoint in Table 5-2) were assigned only when there were experimental data

located for the chemical under evaluation. In addition, the experimental data must have been collected from a well conducted study specifically

designed to evaluate the endpoint under review. If the endpoint was estimated using experimental data from a close structural analog, by

professional judgment, or from a computerized model, then the next-level designation was assigned (e.g., use of data from a structural analog

that would yield a designation of very high would result in a designation of high for the chemical in review). One exception is for the estimated

persistence of polymers with an average MW >1,000 daltons, which may result in a Very High designation.

** The details as to how each endpoint was evaluated are described below and in the DfE full criteria document, DfE Alternatives Assessment

Criteria for Hazard Evaluation, available at: http://www2.epa.gov/saferchoice/alternatives-assessment-criteria-hazard-evaluation.

quantitative analysis and health risk assessment of … · 2017-05-18 · tbp-dbpe...

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