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Safety evaluation strategy of non-intentionally added substances

(NIAS) Sander Koster

Outline

• Safety evaluation of FCM

• Definition of non-intentionally added substances (NIAS)

• NIAS safety evaluation strategies

• Novel safety strategy applied to UNKNOWN substances;

an example on a paper FCM

• Challenges for applying TTC to unknowns

• Outlook

Sander Koster

Safety evaluation of FCM

Safety evaluation of FCM: intentionally added substances (IAS)

Traditionally, check composition starting materials FCM (IAS) +

compliance specific migration limit (SML) substances.

Examples of IAS;

Monomers

Pre-polymers

Base materials

Antioxidants

Polymer production aids

Catalysts

…

Sander Koster


No full safety evaluation!

Unknowns?

IAS

Safety evaluation of FCM; IAS + NIAS

Plastics legislation requires IAS and NIAS (non-intentionally added

substances) to be assessed.

article 3 of the Plastic Regulation (EU) No 10/2011 ‘non-

intentionally added substance (NIAS)’; an impurity in the

substances used or a reaction intermediate formed during the

production process or a decomposition or reaction product.

Sander Koster


Sander Koster


Definition NIAS; recitals Plastic Regulation (EU) No 10/2011

(18) Substances used in the manufacture of plastic materials or articles may contain

impurities originating from their manufacturing or

extraction process. These impurities are non-intentionally added together

with the substance in the manufacture of the plastic material (non-intentionally

added substance – NIAS). As far as they are relevant for the risk assessment the

main impurities of a substance should be considered and if

necessary be included in the specifications of a substance. However it is not

possible to list and consider all impurities in the authorisation. Therefore they

may be present in the material or article but not included in the

Union list.

Sander Koster


Definition NIAS; recitals Plastic Regulation (EU) No 10/2011

(20) During the manufacture and use of plastic materials and articles reaction

and degradation products can be formed. These reaction and degradation

products are non-intentionally present in the plastic material (NIAS). As far as they are

relevant for the risk assessment the main reaction and degradation products of the intended

application of a substance should be considered and included in the restrictions of the

substance. However it is not possible to list and consider all reaction and degradation

products in the authorisation. Therefore they should not be listed as single entries in the

Union list. Any potential health risk in the final material or article arising

from reaction and degradation products should be assessed by the

manufacturer in accordance with internationally recognised scientific

principles on risk assessment.

Examples NIAS

Example of NIAS;

Oxidation product e.g. oxidized irgafos 168

Oligomers?

Impurities

Contaminants of processing/transportation

Starting substances to make e.g. bisphenol-A (acetone, phenol)

…

Does it matter that definition of NIAS can be interpreted differently?

No Final article should not endanger human health

Yes - If detected peak is NIAS it may be present in article

- If detected peak is IAS it should be petitioned.

task ILSI working group on NIAS

Sander Koster


How to do your safety evaluation for NIAS?

Applying traditional approach requires that EVERYTHING that

migrates is identified and risk assessed (full composition, IAS+NIAS).

Obtain information on NIAS throughout the value chain?

Feasible?

What to do if full identification is not possible…?

Sander Koster


Introduction

Most relevant data to start with:

Identity (purity, size, shape, surface area, etc…)

Physico-chemical properties (chemical reactivity, (photo-) catalytic

reactivity, surface charge, etc…)

Changes in either identity and/or physico-chemical properties may

introduce specific hazards

rt rt

Current approach Innovation

Focus on full identification

• Identify & quantify all components

• Hazard & risk assessment for each

individual component

• Low detection levels required

• CRM substances < 10 ppb not covered

resp

on

se

10 ppb

Exposure threshold

Focus on toxicological relevance

• Threshold for identification uprated from

10 ppb to 90 µg exposure per day

• Hazard & risk assessment only

for substances above exposure threshold

• High risk compounds covered by

targeted analyses

Innovation concept for safety evaluation of NIAS

Sander Koster


Innovation concept for safety evaluation of FCM

Generic health limit based on the TTC decision tree

(Kroes et al 2004)

Basic principle in toxicology:

" Everything is poison and there is poison

in everything. It is the dose that makes a

thing a poison."

Risk = Exposure x Hazard

Paracelcus father of the toxicology

Sander Koster


Response

rt

Intake

STEP 1 Translate response into intake and identify peaks

corresponding with intakes of more than 90 µg/p/d

STEP 3, exclude: (structural alerts for) genotoxicity

STEP 2, exclude: proteins (or assess safety)

non-essential/heavy metals

metal containing compounds

dioxin-like chemicals

high potent genotoxic compounds

organophosphates

STEP 5, assess allergenicity

Exposure threshold1

STEP 4 Identify and assess

compounds with

intakes >90 µg/p/d

and non-excluded

compounds

Rennen et al.

2011

1 based on Cramer class III

Sander Koster


Step 1: Screening techniques

Combination of techniques covering broad spectrum of

substances

• Headspace/SPME GC-MS Volatile substances

• GC-FID/MS Semi-volatile subst.

Medium polar/apolar subst.

• Derivatisation* GC-FID/MS Non/semi-volatile subst.

Small polar/medium polar

subst.

• LC-UV/light scattering/MS Non volatile subst.

Polar – apolar subst.

*silylation makes non-volatile substances more volatile

Sander Koster


Step 1: Example NIAS; semi volatiles (GC-MS)

90 µg daily intake

(line to guide the eye)

Tenax

10 days 60 °C

GC-MS,

AT-5 column

1 0 . 0 0 1 5 . 0 0 2 0 . 0 0 2 5 . 0 0 3 0 . 0 0 3 5 . 0 0 4 0 . 0 0 4 5 . 0 0

5 0 0 0 0

1 0 0 0 0 0

1 5 0 0 0 0

2 0 0 0 0 0

2 5 0 0 0 0

3 0 0 0 0 0

3 5 0 0 0 0

4 0 0 0 0 0

4 5 0 0 0 0

5 0 0 0 0 0

5 5 0 0 0 0

T i m e - - >

A b u n d a n c e

T I C : p p n d _ 0 2 3 . D \ d a t a . m s

IS

10 ppb

(line to guide the eye)

Note;

check compliance IAS+ use

chemical information to predict NIAS

Sander Koster


Sander Koster


Step 1: Example NIAS; non-volatiles (LC-ELSD/UV)

mV

-17.40

-17.20

-17.00

-16.80

-16.60

-16.40

-16.20

-16.00

-15.80

-15.60

-15.40

-15.20

-15.00

-14.80

-14.60

-14.40

-14.20

-14.00

Minutes

5.00 10.00 15.00 20.00 25.00 30.00 35.00 40.00 45.00 50.00 55.00

Tenax

10 days 60 °C

H2O ACN,

RP column

90 µg daily intake

IS

Step 2: exclude known high toxic compounds and other TTC excluded classes

• Aflatoxins (relevant for material of biological origin)

• Azoxy substances (not likely in FCM)

• N-nitroso compounds (in rubber materials)

• Steroids (negligible in FCM)

• Dioxins (biological origin + chlorine containing FCM)

• High MW substances e.g. polymers (in general not of tox relevance)

• Non-essential metals (targeted analysis: ICP MS)

• Proteins (screen for allergenicity, see step 5)

• Organophosphates (targeted analysis;TTC of 18 µg/p/d)

Not all classes relevant for FCM. Exclusion based on available

information; expert judgment and/or targeted analysis

Excluded by analysis - ok

ok

ok

ok

To do

Excluded by expert judgment - ok

ok

ok

ok

ok

ok

Sander Koster


Sander Koster


Step 2: Example NIAS; organophosphates (GC-NPD)

NB -pesticides excluded with targeted LC-MS

-LC-MS accurate mass to exclude P

18 µg daily intake

blank

blank

Step 3: Exclude (structural alerts for) genotoxicity

Chemical analysis

Excluding genotoxicity by chemical analysis very difficult (21 structural

alerts).

Bioassays

Conventional assays

AMES, MLA, CA not developed for complex matrices

(higher concentration sensitivity required)

New developments; e.g. Bluescreen

• Luminescent assay (sensitive)

• Human cell line covering endpoints of Ames, micronucleus and

mouse lymphoma assay and cytotoxicity

• GADD45a gene

• Assay validated for pharmaceutical formulations

• High throughput! (96 well-format)

• Test protocol developed for complex matrices

Sander Koster


Step 4: Evaluation of compounds above 90 µg daily exposure

• Identification and quantification of substances > 90µg daily exposure

• Determine substance specific threshold

• Refinement Cramer classification using:

Cramer (1978)

open source applications such as Toxtree

• Substance specific toxicity data

retrieved from public literature

toxicity data from structural related substances (read-

across).

Risk assessment; from ~60 substances (FOP) to ~10 substances (TTC)

Sander Koster


Step 5: Assess allergenicity

• Proteins might give allergic responses in sensitive people and

should therefore be evaluated

• FCMs with ingredients of biological origin might contain proteins

• If considered relevant screening for known allergens and specific

risk assessment

• Here assumed that allergens are not part of the FCM under

investigation

Sander Koster


Challenges for applying TTC approach to unknowns?

Combination toxicity – dose/concentration addition

Current analytical approaches do not take this into account.

However, at low exposure …

• Synergistic effects only when 2 or more compounds are above

effect level (not likely at low TTC exposure)

• Cumulative effects is depending on potency, limited effect to be

expected at low (90 µg/p/d) exposure. Eg.:

• 21 or 26 organophosphates all at 90 µg/p/d => 360 or 250 µg/p/d

when potency corrected for acute or chronic effects, respectively

• 7 or 11 triazoles all at 90 µg/p/d => 240 or 300 µg/p/d when

potency corrected for acute or chronic effects, respectively

Leeman et al. in preparation

Sander Koster



Bio-accumulating substances

Log Po/w as ‘marker’ for accumulation

Three studies where no relation was found between log Po/w and

NOAEL:

• Ravenzwaay (2011): 111 NOAELs from developmental rat studies

(Log Po/w: -4.3 to 15; median 2.12)

• Ravenzwaay (2012): 104 NOAELs from developmental rabbit

studies (Log Po/w: -13 to 15)

• Kalkhof (2012): 824 NOAELs from (28/90 day) repeated dose

studies (Log Po/w: -2.76 to 7.1 [5th/95th Percentile]; median 2.36)

• Relevance of accumulation at low exposure???

(polyhalogenated and metals already excluded)

Sander Koster


Universal response detectors used?

-GC-FID/MS proven to give ‘uniform’ response for different

analytes (factor ~6)

-Light scattering detectors (e.g. ELSD) for LC have fair

uniformity (factor ~6) when operated with post-column addition

of reverse gradient

Do you detect all substance in extracts?

No technique exists that can do so, however state-of-the-art

used here.

Sander Koster



TTC should not be used for mixtures containing unknown chemical

structures?

Combination toxicity effects marginal?

Analytical screening as good as other approaches?

What more arguments needed?

Sander Koster



Outlook

• Efficient evaluation of NIAS requires novel approaches.

• Innovation concept for safety evaluation is pragmatic and feasible.

• Increase analytical screening from 10 ppb to 90 µg daily exposure

• Challenges

• Information transfer through value chain

• Analysis

• Relevant exposure data (FACET?)

• Bioaccumulation, combination toxicity

• Applying to mixtures of unknown chemical structures

• Sensitivity genotoxicity assays

• EFSA

• More demonstrators needed!

• ILSI guidance on NIAS in preparation.

Sander Koster


Thank you for your attention!

See you in autumn 2013 (?) at ILSI! For more information: [email protected]

TNO, The Netherlands: www.tno.nl/foodsafety

Sander Koster


Acknowledgements

Lisette Krul

Winfried Leeman

Monique Rennen

Geert Houben

ILSI and members of ILSI WG on NIAS

mailto:[email protected]

http://www.tno.nl/foodsafety

TTC decision tree; what’s in it?

Exclude:

-proteins

-non-essential/heavy metals

-steroids

-metal containing compounds

-dioxin-like chemicals

-high molecular weight substances

Exclude:

high potent genotoxic compounds

-aflatoxin-like chemicals

-azoxy compounds

-N-nitroso compounds

TTC: 0.15 microgram/p/d Exclude:

Structural alerts for genotoxicity

TTC: 18 microgram/p/d Exclude:

Organophosphates

TTC: 90 microgram/p/d

excluded

excluded

excluded

excluded

present

present

present

Substance specific risk assessment/

Legal limits

Substance specific risk assessment/

Legal limits

Sander Koster


present

LC-ELSD

Sander Koster


Sorb

itol -

2.7

37

Meth

ionin

e-1

- 3

.006

Meth

ionin

e-2

- 3

.988

Quin

ine -

15.7

20

16.0

44

Sulp

ham

eth

azon -

17.6

05

Bis

phenol-A

- 2

6.4

93

17-O

H P

rogeste

ron -

29.1

60

Tebufe

nozid

- 3

2.5

56

mV

-18.00

-16.00

-14.00

-12.00

-10.00

-8.00

-6.00

-4.00

-2.00

0.00

2.00

4.00

6.00

8.00

Minutes

0.00 2.00 4.00 6.00 8.00 10.00 12.00 14.00 16.00 18.00 20.00 22.00 24.00 26.00 28.00 30.00 32.00 34.00 36.00 38.00 40.00 42.00 44.00 46.00 48.00 50.00

Sorb

itol -

2.7

42

Meth

ionin

e-1

- 3

.004

Meth

ionin

e-2

- 3

.991

Quin

ine -

15.7

07

Sulp

ham

eth

azon -

17.5

99

Bis

phenol-A

- 2

6.4

86

17-O

H P

rogeste

ron -

29.1

60

Tebufe

nozid

- 3

2.5

61

mV

-18.00

-16.00

-14.00

-12.00

-10.00

-8.00

-6.00

-4.00

-2.00

0.00

2.00

4.00

6.00

8.00

10.00

12.00

Minutes

0.00 2.00 4.00 6.00 8.00 10.00 12.00 14.00 16.00 18.00 20.00 22.00 24.00 26.00 28.00 30.00 32.00 34.00 36.00 38.00 40.00 42.00 44.00 46.00 48.00 50.00

Normal

gradient

Reversed

gradient

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