Detection of nanoparticles in food - an analytical challenge Stefan Weigel, Ruud Peters IFT International Food Nanoscience Conference July 17, 2010 Chicago, IL, USA
Nano and food: a reality Naturally occuring nano-particles milk: casein micelles (~ 100 nm), whey proteins (~ 3 nm) Conventionally produced food Mayonnaise, Sauce Béarnaise: nano-sized droplets
Nano and food: a reality Engineered nano-particles (ENP) nano-particles in PET bottles (e.g. titanium nitride, nano-clay) nano-silver in food containers nano-encapsulated ingredients/preservatives and, and, and...
Main application areas Food packaging Sales packaging nano-particles incorporated in polymer matrix: clay, TiN, nylon, TiO 2, ZnO,... improved mechanical and barrier properties, UV-protection Active packaging nano-silver incorporated/coated: antimicrobial activity absorbers, scavengers Preservation of quality, extension of shelf life
Main application areas Food ingredients/supplements/auxiliaries Approved additives: SiO 2,TiO 2 (nano-fraction?) Nano-encapsulates preservatives, vitamins, antioxidants,... increased solubility, oxidation protection Nano-formulated ingredients e.g. micelles improved texture, stabilty of emulsions,...
Nano-Food: Risk assessment The European Food Safety Authority (EFSA): The Scientific Committee makes a series of recommendations; in particular, actions should be taken to develop methods to detect and measure ENMs [engineered nanomaterials] in food/feed and biological tissues, to survey the use of ENMs in the food/feed area, to assess the exposure in consumers and livestock, and to generate information on the toxicity of different ENMs. (Scientific Opinion of the Scientific Committee on a request from the European Commission on the Potential Risks Arising from Nanoscience and Nanotechnologies on Food and Feed Safety. The EFSA Journal (2009) 958, 1-39)
Analytical methods: Challenges Methods available for pure NP, only few for complex matrices Detection levels: ppm to ppb levels Availability of standard materials Diversity of ENP Ag
Analytical methods: Challenges Interaction of ENP with matrix, behaviour largely unknown Aggregation, agglomeration, affinity for surfaces Stability of ENP (dissolution, enzymatic digestion,...) Natural NP present in food biogenic geogenic
Analytical methods: Overview Potentially applicable ATOFMS ATOFMS ATOFMS ATOFMS EEM EEM EEM EEM BET BET BET BET AEM AEM AEM AEM CE CE CE CE AES AES AES AES CPC CPC CPC CPC DMA DMA DMA DMA CFUF CFUF CFUF CFUF DLS DLS DLS DLS TEM TEM TEM TEM PCS PCS PCS PCS QELS QELS QELS QELS EM EM EM EM EDS EDS EDS EDS HDC HDC HDC HDC ICP ICP ICP ICP-MS MS MS MS AFM AFM AFM AFM SEM SEM SEM SEM NMR NMR NMR NMR NSOM NSOM NSOM NSOM LIBD LIBD LIBD LIBD HPLC HPLC HPLC HPLC FCS FCS FCS FCS FFF FFF FFF FFF WetSEM WetSEM WetSEM WetSEM UV UV UV UV TOF TOF TOF TOF-MS MS MS MS TGA TGA TGA TGA SMPS SMPS SMPS SMPS SLS SLS SLS SLS SANS SANS SANS SANS SEC SEC SEC SEC XPS XPS XPS XPS XRM XRM XRM XRM XRF XRF XRF XRF XRD XRD XRD XRD XAS XAS XAS XAS
Analytical methods: Selection Food safety approach (statutory testing) presence identity: chemical composition, size (distribution) concentration (mass, number) Imaging (in matrix), screening rapid, high throughput, cost efficient, robust Confirmatory methods unambigious identification, quantification
Analytical methods: Imaging Transmission Electron Microscopy (TEM) Coffee with coffee creamer (containing silica - E551) oil Dilution 2 x water 10 µl on carbon coated copper grids, dried at room temp. oil TEM (JEOL, JEM-1011) SiO2 152 nm 67 nm oil
Analytical methods: Imaging Environmental scanning electron microscopy (ESEM) Energy dispersion spectroscopy (EDS) Bread sample from Sicily Bread sample from Malaysia Gatti et al., Crit Rev Food Sci Nutr 49, 275-282 (2009)
Analytical methods: Confirmatory Sample preparation extraction of ENP from sample matrix removal of interfering matrix components enrichment of target analytes Analytical separation/fractionation by size by physico-chemical properties Detection identification, characterisation quantification
Sample preparation Digestion of matrix chemical (hydrolysis, oxidation, reduction) enzymatic thermal Removal of matrix, pre-concentration, pre-fractionation filtration techniques (e.g. ultra-, membrane tangential flow filtration ) sedimentation, centrifugation ultrasonic extraction solvent extraction (LLE)
Analytical separation Field flow fractionation (FFF) wide dynamic range (~ 1 1000 nm) varioius options available optimalisation of membrane and eluent needed from: Hassellöv et al., Ecotoxicology (2008) 17:344-361
Analytical separation Hydrodynamic chromatography separation by size dependant exclusion from wall, diffusion in laminar flow wide dynamic range (<10 bis >1000 nm) from: Blom et al., Anal. Chem. (2003) 75:6761-6768
Analytical separation Size exclusion chromatography (gel permeation chr.) diffusion into restricted access materials range <1 10 nm, depending on material good separation/resolution HPLC separation by physico-chemical properties very good separation suitable for small particles, e.g. fullerenes
Detection Light scattering determination particle size, size distribution various options: SLS, DLS, MALS,... size range ca. 1 bis > 1000 nm
Detection UV-Detector/DAD Fluorescence detector (FLD) Mass spectrometry (MS) ICP-MS: for inorganic NP, elemental composition desorption-ms, e.g. MALDI-ToF-MS electrospray quadrupol-ms secondary ion-ms (SIMS)... ICP-MS
Example: Nano-silver 1g sample + 2 ml H 2 O, homogenisation Chemical oxidative digestion Filtration (0.45 µm), centrifugation Resuspension in 1 ml eluent (0.1 mm SDS) HDC-ICP-MS
Example: Nano-silver Results Strawberry spiked 0.1 mg/kg
Example: Nano-silver Results: Validation LOQ Range Recovery n=6 RSD Water 20 µg/kg 0.02-2 mg/kg 84 12 % Apple juice 20 µg/kg 0.02-2 mg/kg 86 23 % Tomato 20 µg/kg 0.02-2 mg/kg 85 22 % Strawberry 20 µg/kg 0.02-2 mg/kg 66 15 %
Example: Silica in food Silica is an approved food additive (E551) Used as anti-caking agent in food industry E551 is added to powdered products, e.g. spices, soups, pancake mixtures etc.
Example: Silica in food Does E551 contain a nano-sized fraction? suspension in water TEM agglomerates (200-1000 nm) HDC-ICP ICP-MS
Example: Silica in food Effects in real samples coffee creamer containing E551 ~ 150 nm ~ 20 nm creamer in cold water (20 C) creamer in hot coffee (70 C)
Analytical methods: The pitfalls... Temperature dependance Matrix effects SiO 2 signal strongly depending on matrix - pancake mix reduces silica signal - pudding powder does not... Impact on size of meso-silver - increase of nano-fraction upon addition of matrix - shift of size distribution
Conclusions Electron microscopy is a suitable tool for the imaging of nanoparticles in complex matrices HDC-ICP-MS is a viable technique for the analysis of inorganic NP Interaction of NP with matrix needs to be adressed Sample preparation is a key issue Standards and reference materials needed, well characterised and stable
European collaborative research project Validated methods for ENP in food matrix Inorganic NP: silver, silica, TiO 2,... Organic NP: encapsulates Reference materials for ENP in food Start: January 2010, duration 3 years www.nanolyse.eu (launch August 2010)
Thanks for your attention contact: stefan.weigel@wur.nl Wageningen UR IFT International Food Nanoscience Conference July 17, 2010 Chicago, IL, USA