Three major liquid crystal manufacturers (Merck, Chisso, and Dainippon Ink) committed to not introducing acutely toxic or mutagenic liquid crystal materials to the market. They conduct intensive toxicological and ecotoxicological investigations on liquid crystals according to international guidelines and legal regulations. Test results found that commercial liquid crystals are not acutely toxic, mutagenic, or harmful to aquatic organisms, though they are not readily biodegradable. Overall, comprehensive investigations concluded that no marketed liquid crystals pose toxicity risks.

1. Safety of LC Materials: Toxicological and
Eco-Toxicological Investigations
Dr. Werner Becker, Liquid Crystals Division / Technology & Regulatory Affairs
MDT Workshop “Responsible Care and Legal Compliance”, 15th Oct. 2008

2. Presentation Outline
• Self-commitment and statement of the 3 major
LC manufacturers
• Design of toxicological studies and test
strategy for liquid crystals
• Short term toxicity of LCs
• Mutagenicity of LCs
• Ecotoxicological properties of LCs
• Conclusions
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Dr. Werner Becker, Merck KGaA, LC T & RA Safety of LC Materials: Toxicological and Ecotoxicol. Investigations MDT Workshop 15th Oct. 2008

3. Self-Commitment
Merck KGaA has committed itself to not
introduce into the market liquid crystal materials
which are either acutely toxic or mutagenic.
Likewise Chisso Corp. and Dainippon Ink have
committed themselves, too.
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Dr. Werner Becker, Merck KGaA, LC T & RA Safety of LC Materials: Toxicological and Ecotoxicol. Investigations MDT Workshop 15th Oct. 2008

4. Statement on the Safety of
Liquid Crystal Materials
We, the three liquid crystal material suppliers: Merck, Chisso and Dainippon Ink
maintain the safety of our products, both towards the users and the environment.
Therefore, we carry out intensive toxicological investigations and do not introduce
the acutely toxic or mutagenic liquid crystal materials into the market.
Moreover, we investigate the ecotoxicological properties of our liquid crystal
materials in compliance with legal regulations and as precautionary measures
according to the principles of " Responsible Care " and " Product Stewardship ".
Merck KGaA: Chisso Corporation: Dainippon Ink and Chemicals:
Mr. Walter Galinat Dr. Yasuyuki Goto, Dr. Haruyoshi Takatsu,
Executive Vice President Executive Officer & General Manager, R&D
Liquid Crystals Division General Manager,
Liquid Crystals Division
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Dr. Werner Becker, Merck KGaA, LC T & RA Safety of LC Materials: Toxicological and Ecotoxicol. Investigations MDT Workshop 15th Oct. 2008

5. Design of the Toxicological Studies
Studies are performed:
• during the development of the liquid crystal compounds (i.e.
before introduction into the market), bacteria mutagenicity even
before sampling to customers
• without legal obligation
• according to international guidelines (OECD, EU, MITI)
• in compliance with Good Laboratory Practice-Standards
• following the European and national regulations for animal welfare
• including analogy considerations (Homologous Series Strategy)
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Dr. Werner Becker, Merck KGaA, LC T & RA Safety of LC Materials: Toxicological and Ecotoxicol. Investigations MDT Workshop 15th Oct. 2008

6. Homologous Series Strategy
Homologous series of liquid crystals (CCP-n0CF3):
H
OCF3
H
alkyl side chain core structure terminal
substituent
• Homologous compounds have:
- the same core structure and terminal substituent
- only differ in the length of the alkyl side chain
• Homologue with shortest chain length exhibits most pronounced toxicity.
⇒ Toxicity studies are performed with homologue with shortest side chain.
⇒ These results can be applied to other homologues of the series.
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Dr. Werner Becker, Merck KGaA, LC T & RA Safety of LC Materials: Toxicological and Ecotoxicol. Investigations MDT Workshop 15th Oct. 2008

7. Institute of Toxicology, Darmstadt
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Dr. Werner Becker, Merck KGaA, LC T & RA Safety of LC Materials: Toxicological and Ecotoxicol. Investigations MDT Workshop 15th Oct. 2008

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Dr. Werner Becker, Merck KGaA, LC T & RA Safety of LC Materials: Toxicological and Ecotoxicol. Investigations MDT Workshop 15th Oct. 2008

9. Presentation Outline
• Self-commitment and statement of the 3 major
LC manufacturers
• Design of toxicological studies and test
strategy for liquid crystals
• Short term toxicity of LCs
• Mutagenicity of LCs
• Ecotoxicological properties of LCs
• Conclusions
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Dr. Werner Becker, Merck KGaA, LC T & RA Safety of LC Materials: Toxicological and Ecotoxicol. Investigations MDT Workshop 15th Oct. 2008

10. Acute Toxicity Testing
(Acute Toxic Class Method OECD 423)
• Test species: rats (3 per sex and dose)
• Route of administration: oral
• Dose: max. 2000 mg/kg body weight
• Observation period: 14 days
• Effects recorded: death and all other
signs of toxicity
• Classification and labelling according to the
median lethal dose (LD50)
• LD50: statistical value,
indicates the substance amount at which
50 % of the test animals are considered to die
Test animal: rat
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Dr. Werner Becker, Merck KGaA, LC T & RA Safety of LC Materials: Toxicological and Ecotoxicol. Investigations MDT Workshop 15th Oct. 2008

11. Acute Oral Toxicity of Liquid Crystals
LCs LC-mixtures
No. of substances tested: 269 14
no effect (LD50 > 2000 mg/kg bw) 260 14
harmful (LD50 200 - 2000 mg/kg bw) 8 0
260 LCs and 14 mixtures caused no effects Status:
Sept. 23, 2008
8 LCs are classified as harmful
- only old STN/TN substances concerned, none of the TFT singles
→ concentration in mixtures < 10% LC mixtures are not harmful
1 LC was found to be toxic
→ excluded from further development and not introduced into the market
no marketed LC is acutely toxic and no TFT LC is even harmful !
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Dr. Werner Becker, Merck KGaA, LC T & RA Safety of LC Materials: Toxicological and Ecotoxicol. Investigations MDT Workshop 15th Oct. 2008

12. Presentation Outline
• Self-commitment and statement of the 3 major LC
manufacturers
• Design of toxicological studies and test strategy for
liquid crystals
• Short term toxicity of LCs
• Mutagenicity of LCs
• Bacteria mutagenicity (Ames test)
• Mammalian mutagenicity (Chromosome
Aberration)
• Ecotoxicological properties of LCs
• Conclusions
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Dr. Werner Becker, Merck KGaA, LC T & RA Safety of LC Materials: Toxicological and Ecotoxicol. Investigations MDT Workshop 15th Oct. 2008

13. Mutation: What is a Mutation ?
Cell with nucleus
DNA molecule
Mutagenic substances can cause DNA damage
in the organic purine (guanine, adenine) and
pyrimidine (thymine, cytosine) bases
Mutation is the critical event in the initiation of cancer
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Dr. Werner Becker, Merck KGaA, LC T & RA Safety of LC Materials: Toxicological and Ecotoxicol. Investigations MDT Workshop 15th Oct. 2008

14. Principle of the Bacterial Mutagenicity
Assay (Ames-Test OECD 471, 473)
Mutant bacteria:
Salmonella typhimurium (his-) and/or
Escherichia coli (trp-)
+/- metabolic
activation (S9)
treatment with test substance (TS)
TS is not mutagenic (his-/trp-): TS is mutagenic (his+/trp+):
no backmutation to wildtype bacteria backmutation to wildtype bacteria
no revertant bacterial colonies revertant bacterial colonies
in his-free and/or trp-free media in his-free and/or trp-free media
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Dr. Werner Becker, Merck KGaA, LC T & RA Safety of LC Materials: Toxicological and Ecotoxicol. Investigations MDT Workshop 15th Oct. 2008

15. Mutagenic Potential of LCs in Bacteria
LCs LC-mixtures
No. of LCs studied: 278 14
not mutagenic: 276 14
276 LC compounds and 14 mixtures investigated are
not mutagenic in bacteria
2 LCs were mutagenic in bacteria and were not further
developed and not introduced into the market
no marketed LC is mutagenic ! Status:
Sept. 23, 2008
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Dr. Werner Becker, Merck KGaA, LC T & RA Safety of LC Materials: Toxicological and Ecotoxicol. Investigations MDT Workshop 15th Oct. 2008

16. Principle of the Chromosome
Aberration Test (OECD 476)
Cells from Chinese Hamster Lung (CHL) or Ovary (CHO)
+/- metabolic activation S9
treatment with test substance (TS)
expression period
TS is not clastogenic: TS is clastogenic:
No Chromosome Aberrations Chromosome Aberrations
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Dr. Werner Becker, Merck KGaA, LC T & RA Safety of LC Materials: Toxicological and Ecotoxicol. Investigations MDT Workshop 15th Oct. 2008

17. Clastogenic Potential of LCs in
Mammalian cells
• Several chromosome aberration studies were performed
for notification purposes
Number of LCs studied: 46
Status:
not clastogenic: 46 Sept. 25, 2008
no LC compound is clastogenic in mammalian cells
no indication for a carcinogenic potential of LCs
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Dr. Werner Becker, Merck KGaA, LC T & RA Safety of LC Materials: Toxicological and Ecotoxicol. Investigations MDT Workshop 15th Oct. 2008

18. Presentation Outline
• Self-commitment and statement of the 3 major LC
manufacturers
• Design of toxicological studies and test strategy for
liquid crystals
• Short term toxicity of LCs
• Mutagenicity of LCs
• Ecotoxicological properties of LCs
• Aquatic toxicity (algae, fish, daphnia)
• Biodegradation
• Bioaccumulation (Bioconcentration)
• Conclusions
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Dr. Werner Becker, Merck KGaA, LC T & RA Safety of LC Materials: Toxicological and Ecotoxicol. Investigations MDT Workshop 15th Oct. 2008

19. Aquatic Test Organisms
unicellular green algae
Zebra fish (Desmodesmus subspicatus)
(Danio rerio)
Length: up to 5 cm
Water flea
(Daphnia magna)
Size: up to 5 mm
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Dr. Werner Becker, Merck KGaA, LC T & RA Safety of LC Materials: Toxicological and Ecotoxicol. Investigations MDT Workshop 15th Oct. 2008

20. Aquatic Toxicity of Liquid Crystals
• LCs are poorly water soluble and difficult to test in aquatic systems
• Available tests
- fish: 7 LCs
- daphnia: 39 LCs (+ 1 mixture)
- algae: 13 LCs (+ 1 mixture)
- bacteria: 8 LCs Green algae Daphnia magna
Results
No effects up to the highest recommended test substance concentration
(100 mg/L) for aquatic organisms
Liquid Crystals are not harmful to aquatic organisms !
Status:
Sept. 23, 2008
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Dr. Werner Becker, Merck KGaA, LC T & RA Safety of LC Materials: Toxicological and Ecotoxicol. Investigations MDT Workshop 15th Oct. 2008

21. Biodegradation
Amoeba
Bacteria
Activated sludge
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Dr. Werner Becker, Merck KGaA, LC T & RA Safety of LC Materials: Toxicological and Ecotoxicol. Investigations MDT Workshop 15th Oct. 2008

22. Ready Biodegradability of Liquid Crystals
Liquid crystals (in particular SFM compounds) are chemically
very stable and not readily biodegradable.
Degradation rates were between 0 and 55 % after 28 days.
Ester type substances and CN substituted compounds are partly
biodegradable whereas SFM compounds are not
Substances tested to date : 89
Status:
Sept. 23, 2008
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Dr. Werner Becker, Merck KGaA, LC T & RA Safety of LC Materials: Toxicological and Ecotoxicol. Investigations MDT Workshop 15th Oct. 2008

23. Bioaccumulation (Bioconcentration)
• Guideline: Japanese Guideline (Medaka,
Oryzias latipes)
• Test species: Japanese rice fish
• Duration:
- Uptake phase: 28 days (can be extended to up to 60 days)
- Depuration phase
• Analysis of test substance concentration in water and fish
• Results are given as Bioconcentration factor (BCF)
equilibrium concentration of test substance in fish
BCF =
equilibrium concentration of test substance in water
For notification purposes, 46 LCs were tested for bioaccumulation
Conclusion: Liquid Crystals do not show a significant
bioaccumulation potential !
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Dr. Werner Becker, Merck KGaA, LC T & RA Safety of LC Materials: Toxicological and Ecotoxicol. Investigations MDT Workshop 15th Oct. 2008

24. Summary of Toxicological and Eco-
toxicological Testing Results
Commercial liquid crystal compounds
are not acutely toxic,
are not mutagenic in bacteria (Ames-Test),
are not clastogenic in the chromosome aberration test in
mammalian cells,
are not suspicious of carcinogenicity,
are not harmful to aquatic organisms (bacteria, algae,
daphnia, fish),
do not show a significant bioaccumulation potential,
but are not readily biodegradable.
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Dr. Werner Becker, Merck KGaA, LC T & RA Safety of LC Materials: Toxicological and Ecotoxicol. Investigations MDT Workshop 15th Oct. 2008

25. What is a Poison ?
All things are poison; there is
nothing that is not a poison;
only the dose makes that a
thing is not a poison.
Paracelsus (1493-1541)
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Dr. Werner Becker, Merck KGaA, LC T & RA Safety of LC Materials: Toxicological and Ecotoxicol. Investigations MDT Workshop 15th Oct. 2008

26. Thank you for your attention !
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Dr. Werner Becker, Merck KGaA, LC T & RA Safety of LC Materials: Toxicological and Ecotoxicol. Investigations MDT Workshop 15th Oct. 2008