Screening methods - Scope and limitations - M. Stintz, C. Ullmann, F. Babick NanoDefine Outreach Event Brussels, September 217 NanoDefine is funded by the European Community's Seventh Framework Programme under Grant Agreement-64347
Outline What does screening mean? Screening methods Validation of key methods Outcome and publications
Screening methods What does screening mean? Screening method means methods that are used to detect the presence of a substance or class of substances at the level of interest. These methods have the capability for a high sample throughput and are used to sift large numbers of samples for potential non-compliant results. They are specifically designed to avoid false compliant results. (22/657/EC: Annex 1: 1.35) What does screening mean in this context? Robust and easy implementable method for rapid distinction between nano/non-nano and avoid false positive decisions acc. to the EC recommendation. Potential screening methods: Analytical Centrifugation methods (AC) Dynamic light scattering (DLS) Electric mobility analysis MiniTEM Particle tracking analysis (PTA/NTA) SingIe particle ICP-MS
Screening methods
Analytical Centrifugation Principle Heavier particles sediment faster than smaller ones do: Separation of particles is caused by migration velocity. Achievements Evaluation of limiting factors and practical method improvements on: - Cuvette-type centrifuge - Disk-type centrifuge - Analytical Ultracentrifuge centrifugal force Stokes equation Differential / integral sedimentation analysis principle Data-based comparison of techniques and settings (wavelength, ) Validation of method performances with real world substance (nano/non-nano) extintion cross section [nm²] 1E+8 1E+6 1E+4 1E+2 1E+ Polystyrene / Water Silica / Water 1E-2 Titania / Water 1E-4 PMMA / Water 1 1 1 1 particle size [nm] Local concentration measurement: Material dependent extinction cross section
Centrifugation techniques / instruments LumiSizer 61 & 651 (LUM GmbH) DC 24UHR (CPS Instruments) AUC: Modified XL-I ProteomeLab Version (Beckman Coulter / BASF) Cuvette centrifuge Disc centrifuge Cuvette centrifuge detects light extinction at λ=47 nm / 865 nm detects light extinction at λ=45 nm detects RI increment at λ=67 nm integral sedimentation differential sedimentation integral sedimentation Dresden University of Technology JRC-IHCP BASF & JRC-IHCP
Consistency of centrifugation methods ID-2 Trimodal polystyrene mixture 46 nm/ 1 nm/ 35 nm nominal diameter mixed 1:2:2 by mass (JRC-IRMM) Modal particle size x mod, [nm] Consistency of mode accuracy x mod, 4 3 2 1 Mode 46 nm Mode 1 nm Mode 35 nm - 47 nm 865 nm 45 nm RI nominal cuvette-type AC disc-type AC AUC Number weights [%] Comparison of number quantity 1 8 6 4 2 Mode 46 nm Mode 1 nm Mode 35 nm,4,4 2,7,5,6 16,2 83,4 25,5 74,1 97,3 25,1 74,4 14,4-47 nm 865 nm 45 nm RI nominal cuvette-type AC disc-type AC 85 AUC C. Ullmann et al.: Evaluation of particle sizing by analytical centrifugation using real-world materials for the implementation of the EC definition of the term nanomaterial NanoMetrology France 217 Conference and Exhibition, Paris, 28.-3.6.217 7
Principle Dynamic light scattering Particle s Brownian motion causes fluctuation in scattered light. θ Principle of DLS Particle s size dependent intensity fluctuation norm. correlation funct n g2-1 1..9.8.7.6.5.4.3.2.1. 5 nm 15 nm 22 nm 44 nm increasing size.1.1.1.1 1 1 delay time t, ms Particle s size dependent correlation function Method refinement for non-nano, broadly distributed real world substances: Studies on filtration steps and on delay time impact SEM image of PTFE membrane magnification 25x Improvement of raw data: Correlation function before and after filtration 1. µm PTFE Kaolin measured after membrane filtration (PTFE) C. Ullmann & P. Mueller: SOP, applicability range and method performance description for DLS & MiniTEM. Technical Report D4.6, NanoDefine Consortium, Wageningen, 217
Electric mobility spectroscopy Evaluation of SMPS for NP characterisation Principle: Aerosolisation of suspensions for aerodynamic size determination Advantage: Direct access to number-based size distributions. Prototype development for analysis of extremely small particles by RAMEM: Size range: 1-5nm Principle of DMA particle size [nm] Application of state-of-the-art SMPS for NanoDefine reference materials 4 35 3 25 2 15 1 5 measured mode diameter reference mode diameter High Resolution Mobility Spectrometre prototype by RAMEM Characterisation with mobility standards: - 1.5 nm: Tetraheptyl ammonium Bromide - 1.7 nm: Tetradodecyl ammonium Bromide
MiniTEM Principle Low voltage TEM (25 kv) Advantage: Direct access to number-based size distributions. Achievements Evaluation of method for screening and confirmatory purposes Configuration strategy for the NanoDefine ParticleSizer BF-TEM BF-MiniTEM HAADF- STEM BF-TSEM HAADF-TSEM SE-SEM TableTop -SEM ~1 Comparison of image (Ag-particles) taken by EM-based techniques nm Automated acquisition/ evaluation routine Automated imaging strategies: intelligent sample screening C. Ullmann & P. Mueller: SOP, applicability range and method performance description for DLS & MiniTEM. Technical Report D4.6, NanoDefine Consortium, Wageningen, 217
Particle tracking analysis Principle Size dependent movement of particles (Brownian motion) is captured Advantage: Direct access to number-based size distributions. Schematic of optical configuration Achievements New software release for PTA instruments Particle concentration measurements Calibration and measurement protocol for number concentration with awareness of detection threshold and camera level Uncalibrated / calibrated detection threshold Concentration (ppml) 7 6 5 4 3 2 1 3 4 5 6 Uncalibrated 7 8 9 1 11 12 13 14 Camera Level 3 4 5 6 7 8 9 1 11 12 13 Calibrated 14 Measurements under flow: reduction in variation P. Vincent: NanoSight NTA Concentration Measurement Upgrade Technical note, application note, brochure, 215
Single particle ICP-MS Principle inductively coupled plasma-mass spectrometry (ICP- MS) operated in the single particle mode Advantage: Direct access to number-based size distributions. Achievements Platform independent software using dwell times in the range of 1-1 ms (and higher) Development multi-element spicp.ms method Calculation of particle size distribution and number concentration Autom. determination of key evaluation parameters: detection sensitivity and nebulisation efficiency Statistical evaluation of the recorded data to raise user awareness of artefacts Recommendation of acquisition parameters (dwell time, ) Validation of methods for substances and products Impact of dwell time setting Vaporisation of particles in plasma Signal distribution view of NIST 813
Validation
Validation of key screening methods Intermediate precision of method within 5 days according to ISO 5725-3:1994, uncertainty obtained by ANOVA Individually prepared samples incl. preparation, measurement, data analysis, conversion from Q ext (x) or Q 3 (x) to Q (x) Strong cooperation with JRC-IRMM s 2 day Lab day 1 day 2 day p R 1 R 2 R 3 R 1 R 2 R 3 R 1 R 2 R 3 s 2 r s 2 r s 2 r Method 1: Analytical centrifugation (AC): cuvette-type AC disc-type AC analytical ultracentrifuge BaSO 4 nano grade (12 nm 25 nm) BaSO 4 non-nano grade (8 nm 25 nm) SiO 2 trimodal 3/8/12nm (disc-type AC and AUC only) Method 2: Single particle ICP-MS Coated TiO 2 TiO 2 in susnscreen Al 2 O 3 in toothpaste 14
Intermediate precision of AC Analytical centrifugation with optical detection classifies nano/non-nano on screening level. Precision at real substances < 11% RSD (1σ) except using λ = 865 nm Impact of sample preparation, especially agglomerate destruction Cumulative funct. Q(x) [%] 1 9 8 7 6 5 4 3 2 1 Particle size x [nm] Cuvette-type AC: BaSO 4 nano grade, light source λ= 47 nm Cuvette-type AC: Comparison of results - Illumination at λ = 47 nm & λ = 865 nm 25 3 Rel. std. dev. (1σ) [%] 2 15 1 5 ux1, ux5, ux9, Rel. Std. dev. (1σ) [%] 25 2 15 1 5 47 nm 865 nm repeatability of measuring subsamples (N=1) measurement method repeatability within days (n=3) method repeatability between days (p=5) method method intermediate precision x1 x5 x9 x1 x5 x9 x1 x5 x9 x1 x5 x9 repeatability of measuring subsamples (N=1) method repeatability within days (n=3) method repeatability between days (p=5) method intermediate precision C. Ullmann, F. Babick, R. Koeber, M. Stintz: Performance of analytical centrifugation for the particle size analysis of real-world materials. Power Technology, 217 15
AC method comparison: nano/non-nano BaSO 4 powder nano grade 12 nm 25 nm BaSO 4 powder non-nano grade 8 nm 25 nm Stokes' median diameter [nm] 1 8 6 4 2 47 nm 865 nm45 nm RI RI 47 nm 865 nm45 nm RI RI Cuvette-AC Disc- AC x 5,3 x 5, AUC I AUC II Cuvette-AC Disc- AC x 5,3 x 5, AUC I AUC II Stokes' median diameter [nm] 8 6 4 2 x 5,3 x 5, 47 nm 865 nm 45 nm RI 47 nm 865 nm 45 nm RI Cuvette-AC Disc-AC AUC I Cuvette-AC Disc-AC AUC I x 5,3 x 5, Rel. std. dev. (1σ) [%] 2 1 47 nm 865 nm 45 nm RI RI 47 nm 865 nm 45 nm RI RI Cuvette-AC Disc- AC u x(5,3) AUC I AUC II Cuvette-AC Disc- AC ux 5,3 ux 5, u x(5,) AUC I AUC II Rel. std. dev. (1σ) [%] 2 1 u x(5,3) u x(5,) 47 nm 865 nm 45 nm RI 47 nm 865 nm 45 nm RI Cuvette-AC Disc-AC AUC I Cuvette-AC Disc-AC AUC I ux 5,3 ux 5, C. Ullmann et al.: Evaluation of particle sizing by analytical centrifugation using real-world materials for the implementation of the EC definition of the term nanomaterial NanoMetrology France 217 Conference and Exhibition, Paris, 28.-3.6.217 16
Outcome Improvements of measurement procedure and data interpretation Validation studies: Data base for e-tool and input for standardisation Added value: comparison and validation of intensity-based and mass-based distributions e.g. for risk assessment (particle release, transport, exposure) Publications: peer reviewed papers: 2 (1 submitted) other publications: 5 proceedings: 1 talks and posters: >8
Selected publications D. Mehn, I. Rio, D. Gilliland, M. Kaiser, K. Vilsmeier, P. Schuck, W. Wohlleben: Analytical ultracentrifugation with fixed and ramped speed for nanomaterial identification- a validation report on three materials in two laboratories, special issue. Under review. Quality in Nanoscience NanoImpact Journal C. Ullmann, F. Babick, R. Koeber, M. Stintz: Performance of analytical centrifugation for the particle size analysis of real-world materials. Powder Technology, 217 C. Ullmann, I. Rio, D. Mehn, D. Gilliland, W. Wohlleben, R. Koeber, F. Babick: Evaluation of particle sizing by analytical centrifugation using real-world materials for the implementation of the EC definition of the term nanomaterial. Accepted for publication in Proceedings NanoMetrology France 217 Conference and Exhibition, Paris: Frankreich, 28.-3.6.217 C. Ullmann & P. Mueller: SOP, applicability range and method performance description for DLS & MiniTEM. Technical Report D4.6, NanoDefine Consortium, Wageningen, 217 F. Babick, J Mielke, W. Wohlleben, S. Weigel, V.-D. Hodoroaba: How reliably can a material be classified as a nanomaterial? Available particle sizing techniques at work. J. Nanopart. Res, 216 P. Vincent: NanoSight NTA Concentration Measurement Upgrade Technical note, application note, brochure, 215
Achnowlegments F. Babick, C. Ullmann, L. Hillemann R. Koeber, D. Gilliland, D. Mehn, I. M. Rio-Echevarria, R. Peters, A. Undas, S. Böhme, C. Cascio D. Kutscher W. Wohlleben, P. Müller, M. Kaiser, K. Vilsmeier, P. Schuck P. Dohanyosova, S. Lopez P. Hole, P. Vincent 19
Achnowlegments The research leading to these results has received funding from the European Union s Seventh Framework Programme (FP7/27-213) under grant agreement n 64347 NanoDefine www.nanodefine.eu) Development of an integrated approach based on validated and standardized methods to support the implementation of the EC s recommendation for a definition of nanomaterials 2
www.nanodefine.eu This project has received funding from the European Union s Seventh Programme for research, technological development and demonstration under grant agreement No 64347