Harald F. Krug - NanoDiaRA · Materials Science &Technolog y Materials meet LifeMaterials Meet Life...

Materials Science & Technology

Harald F. Krug Empa, Department Materials Meet Life, St. Gallen, Switzerland

Functionalised clothing for the Monitoring of Body Functions

MedTech Materials for Osteointegration

Coating technology for Nanocomposites and Fibers Flame retardant

Materials & Additives

Material safety

Biopolymere Materials & Coatings

Materials Sci ence & Technolog y

Materials meet Life Materials Meet Life HF Krug, 19.04.2012, NanoDiaRA 3

Link between Medicine and Toxicology

Bayer scientist Felix Hoffmann synthesized aspirin and heroin in the same month, and the company marketed the two drugs together. This advertisement shows Bayer's pre-1904 logo

Thomas Hartung, Johns Hopkins University



Nano for more then 100 Years

The Tumour Tissue is heated up to a maximum of about 49°C and Cells will be destroyed

Dr. A. Jordan, Charité Berlin

Dr. A. Jordan, Charité Berlin

Aminosilane coated Ironoxide Nanoparticles in aqueous Suspension (MagForce®) at a Concentration of 100 mg/ml Injected into the Tumour Tissue

Magnetic Nanoparticles for Tumour Therapy

P. Gröning / Empa / Nanotechnologie

Magnetic Nanoparticles for Drug Delivery

Magnetic Drug Targeting

P. Gröning / Empa / Nanotechnologie

C.L. Haynes (2010): The emerging field of nanotoxicology Anal. Bioanal. Chem 398, 587-588.

2011

http://www.empa.ch/plugin/template/empa/*/113719

http://www.dechema.de/studien-path-1,123212.html

http://ihcp.jrc.ec.europa.eu/our_activities/nanotechnology/nanoreport-10-11/JRC-EASAC-report.pdf



Nanomaterials – the great Uncertainty?

NANO



Three Nanotox-Principles

The Transport-Principle Limbach LK, Wick P, et al. (2007). Environ. Sci. Technol. 41:4158-4163

The Surface-Principle Nel et al. (2006) Science 311: 622-627 Oberdörster et al., (2000) HEI-Report 96

The Material-Principle

Krug & Wick (2011) Angew. Chem. Int. Ed., 50 (6): 1260-1278



Caveoli (Ø=100nm)

Size Matters!



Small Size but very large specific Surface



One Material – different Modifications and …

Carbon many more …… V2O3

ZnO

Pd



(Material)Surface Properties!

Xia et al. (2009) Annu. Rev. Public Health 30: 137-150



- + TiO2 CB CeO2 ZrO2 ZnO AlOOH 0

2

4

6

8 10

15

20

25

IL-8

Sec

retio

n (x

-fold

)

Same Size – Different Materials

0.5, 5 and 25 g/cm2

Ø: TiO2 10-20 nm Carbon Black (CB) 15 nm CeO2 20 nm ZrO2 10-25 nm ZnO 40 nm AlOOH 40 nm

Kuhlbusch T, Krug HF, and Nau K. (2009) NanoCare: health related aspects of nanomaterials. Karlsruhe, St. Gallen, DECHEMA e.V. http://www.nanopartikel.info/files/content/dana/Dokumente/NanoCare/Publikationen/NanoCare_Final_Report.pdf

http://www.nanopartikel.info/files/content/dana/Dokumente/NanoCare/Publikationen/NanoCare_Final_Report.pdf



Zinc-Story – essential or toxic on the Nano-Level? No difference between nano or bulk ZnO particles

Buerki-Thurnherr T, Xiao L, Diener L, Arslan O, Hirsch C, Maeder-Althaus X, Grieder K, Wampfler B, Mathur S, Wick P, and Krug HF (2012). In vitro mechanistic study towards a better understanding of ZnO nanoparticle toxicity. Nanotoxicology, online first

http://www.ncbi.nlm.nih.gov/pubmed/22394310?dopt=Citation





Variables Associated with Nanomaterial Samples

Variables Associated with Toxicity Test

Variables Associated with Biological Sample

Sample Purification to Remove Possible Biological Interferents

NIST/USA

Depending on desired endpoint, there is a wide variety of

toxicology tests each with their own set of variables

Cell Line: - Choice of Cell Line - Cell Line Identification - Age and Storage of Cell Line - Number of Passages - etc.

Sample Characterization of Raw Nanomaterial (in manufactured form): - Composition and Purity - Size - Shape - Agglomeration etc.

Sample Characterization of Raw Nanomaterial to Determine Biological Impurities - Endotoxin etc.

Dispersion in Biological Media under Appropriate Conditions for Simulated Route of Exposure: - Temperature - Humidity - Gas Concentration - Salinity etc.

Sample Characterization in Biological Media: - Size & Shape - Agglomeration - Protein coating etc.

Set of appropriate positive and negative controls to determine possible interferences with test

etc.

Control of Conditions under which Experiments are Performed: - Temperature - Humidity - Gas Concentration - Salinity - etc.

Variables Associated with in vitro Toxicity Testing (Locascio et al., 2011 in: Nanotechnology Standards Nanostructure Science and Technology, pp: 179-208, Springer



Pulskamp, K., Diabaté, S., Krug, H.F. (2007) Toxicol. Lett. 168: 58-74

The Paradox of Viability Assays

MTT-Assay

WST-Assay



Our findings strongly suggest verifying cytotoxicity data with at least two or more independent test systems for this new class of materials (nanomaterials). Moreover, we intensely recommend standardizing nanotoxicological assays with regard to the material used: there is a clear need for reference materials.

The Paradox of Viability Assays



Fullerene Paradox: Radical Scavenger



Fullerene Paradox: Oxidative Stress



C60 Fluka

Fullerene Paradox: Small Balls (C60) – Big Pitfalls

transfer THF/water

in THF

C60 in THF susp.

Suspension

start

after weeks

in water

Toxic Effect No Effect



C60 Fluka

Fullerene Paradox: Small Balls (C60) – Big Pitfalls

transfer THF/water

in THF

C60 in THF susp.

Suspension

start

after weeks

in water

Toxic Effect No Effect

Spohn P et al. (2009) Environ. Pollut. 157:1134-1139



Wrong Methods/Missing Controls How to judge an effect

Yang H, Liu C, Yang D, Zhang H, Xi Z (2009) J. Appl. Toxicol. 29:69-78



More Pitfalls and Flaws

Toxicologist Non-Toxicologist



High Dosage/ Concentrations



Excessive Delivery of Nanostructured Matter to Submersed Cells …….

Schematic illustrations of the conceivable interaction of insoluble particles with submersed cells grown at the bottom of a well, filled with an appropriate medium of height h. (A) Previously employed picture, (B) more appropriate concept discussed in this study. The number of particles in (A) and (B) is the same.

Wittmaack, K. (2011) ACS Nano, 5: 3766-3778

TiO2(80) particles (27 μg/mL)

At the quoted heights (h) of the media, the mass concentration converts to maximum achievable areal densities of (A-C) 8.4 μg/cm2 and (D-J) 25.2 μg/cm2 (corresponding thicknesses 21.5 and 64.5 nm)



0.5 mg/rat lung: no evidence for carcinogenicity

A minimum of 5 X 3 mg/rat lung, maximum 20 x 6 mg/rat lung by instillation carcinogenicity!



Calculated Overload

Rat lung weight: 0.9 g

Macrophage count: 1.200.000 0,5mg/lung 416 pg / cell: no effects (Yokohira et al., 2009)

15mg/lung 12.500 pg / cell

120mg/lung 100.000 pg / cell tumour induction (Roller, 2009)

3mg/lung 2.500 pg / cell reflects overload conditions



SOPs



Willbe available soon at www.nanoobjects.info and www.nanosafetycluster.eu

http://www.nanoobjects.info/

http://www.nanosafetycluster.eu/



Harmonised(Standardised) Methods

V Viability

I Inflammation

G Genotoxicity

O Oxidative stress



www.nanoobjects.info www.nanopartikel.info

http://www.nanoobjects.info/

http://www.nanopartikel.info/



Exposure & Environment Foreign Entities vs. Endogenous Entities Don’t Believe a Single Method Method Reliability In Vitro in Vivo Correlation Nano-Specific Signatures REACH Challenges for Regulatory Activities Fibre Paradigm Revisited Crowns/Coronas out of Proteins, Lipids and …? How to define “Biopersistence”?

Future Challenges in Nanotoxicology



Future Needs of Nanotoxicology

The huge amount of new chemicals and expected new nanomaterials on the market raises the question for alternative methods for a first screening

In vitro methods should be reliable, robust, sensitive and predictive

Without a responsible process of marketing “nanotechnologies” will fail within the next decade

Support of (nano)toxicological activities/projects is continuously needed; risk characterisation and assessment is crucial

Comparability of Tox-studies is mandatory and should be achieved by use of standardised methods, reference materials and the appropriate controls in each experimental set-up



HF Krug, 16.09.2010, NANO2010 Rom

KIT (NanoCare & DaNa): K. Nau

Materials-Biology Interactions (Empa): A. Bruinink T. Bürki (Thurnherr) M. Gasser C. Hirsch J.-P. Kaiser B. Kopf S. Lischer K. Maniura M. Rösslein M. Rottmar A. Schipanski P. Wick

Cooperations (Selection): NIST, USA NIMS / NIHS, JAPAN Karolinska Institute, Sweden KRISS, Korea

(Nano)Materials: S. Balaban – INT-CFN, Karlsruhe C. Feldmann – Uni Karlsruhe-CFN F. Hennrich – INT-CFN, Karlsruhe H. Hofmann – EPFL, Lausanne FHI Berlin University of Cologne, Germany University of Memphis, USA Bayer AG

Alumni (Nanotox): L. Belyanskaya P. Spohn K. Fischer J. Wörle-Knirsch

Funding: BMBF BAG/BAfU CCMX NCCR EU Industry KTI SNF

Harald F. Krug - NanoDiaRA · Materials Science &Technolog y Materials meet LifeMaterials Meet Life...

Documents

Transcript of Harald F. Krug - NanoDiaRA · Materials Science &Technolog y Materials meet LifeMaterials Meet Life...