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NANOPARTICLES IN FOOD BIOSENSING

NANOPARTICLES IN FOOD BIOSENSING. J.M. Pingarrón* L. Agüí and P. Yáñez-Sedeño Department of Analytical Chemistry. Faculty of Chemistry University Complutense of Madrid 28040-Madrid SPAIN pingarro@quim.ucm.es. NANOJASP’2010 Barcelona, December 2010.

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NANOPARTICLES IN FOOD BIOSENSING

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  1. NANOPARTICLES IN FOOD BIOSENSING J.M. Pingarrón* L. Agüí and P. Yáñez-Sedeño Department of Analytical Chemistry. Faculty of Chemistry University Complutense of Madrid 28040-Madrid SPAIN pingarro@quim.ucm.es NANOJASP’2010 Barcelona, December 2010

  2. Preparation of nanostructured electrode surfaces Research line combining: Advances in sensors technology Development of several (bio)assay-transductor strategies Applications of nanotechnology - Wide range of approaches - Use or not of biological systems Products, processes and systems operating in nanometric magnitude

  3. Advantages Sensitivity Selectivity Repeatability Nanostructured electrode surfaces Improved charge transfer reactions Electrocatalytic ability: lower detection potential Antifouling capability

  4. Au Au Au ELECTROCHEMICAL BIOSENSORS BASED ON GOLD NANOPARTICLE-MODIFIED ELECTRODES Gold nanoparticles Ability to provide a stable surface for biomolecules immobilization retaining their biological activity Permit direct electron-transfer between redox proteins and bulk electrode materials no need for electron-transfer mediators High surface-to-volume ratio High surface energy Ability to decrease the distance between proteins and metal particles Functioning as electron- conducting pathways between the prosthetic groups and the electrode surface Useful interfaces for electrocatalysis of redox processes of H2O2 or NADH Electrochim. Acta., 53 (2008) 5848

  5. ELECTROCHEMICAL BIOSENSORS FOR FOOD ANALYSIS DEVELOPMENT AND INNOVATION FOOD SAFETY FOOD QUALITY EFFICIENT TRACEABILITY SYSTEMS Development of detection, analysis and diagnosis methods  Rapid  Sensitive  Automated screening

  6. AMPEROMETRIC BIOSENSOR FOR HYPOXANTHINE BASED ON IMMOBILIZED XOD ON NANOCRYSTAL GOLD-CARBON PASTE ELECTRODES Gold nanoparticle preparation:Electrodeposition from a HAuCl4 solution on the bulk electrode material • Hypoxanthine (Hx) is formed as a product of nucleotide catabolism during the degradation processes in foodstuffs of animal origin. • Hx is accumulated mostly in the animal muscle and its levels are used as an index of fish and meat freshness in the food industry Determination of hypoxanthine based on the enzyme reaction catalyzed by XOD XOD Hx + O2→ X + H2O2 XOD X + O2 → Uric acid + H2O2

  7. AMPEROMETRIC BIOSENSOR FOR HYPOXANTHINE BASED ON IMMOBILIZED XOD ON NANOCRYSTAL GOLD-CARBON PASTE ELECTRODES LOD at 0.00 V: 2.2x10-7 mol L-1 Kmapp = 18x10-6 mol L-1 Useful lifetime = at least 15 days Sens. Actuators B. 113 (2006) 272 SEM of a GA-BSA-XOD-nAu-CPE biosensor Determination of hypoxanthine in sardines and chicken meat using the GA-BSA-XOD-nAu-CPE biosensor Sample Sardines Chicken Recovery (%) Added (mg/100g) Found (mg/100g) Recovery (%) Added (mg/100g) Found (mg/100g) Non-spiked 1 - 91.7 - - 145 - 2 - 138 - - 85.0 - - 94.7 - - 152 - 3 Spiked 1 - 225 105 - 157.2 103 2 69.6 213 103 60.7 145.1 99 - 210 95 - 160.2 103 3 Mean recoveries ( = 0.05): 101 ± 8 % sardines 102±3% chicken meat

  8. Bienzyme amperometric biosensor using gold nanoparticles-modified electrodes for the determination of inulin in foods Anal. Biochem., 375 (2008) 345-353

  9. DETERMINATION METHODS HPLC UV, RI, ED 1st enzyme biosensor This work INULIN Prebiotic ingredient added to functional foods Vegetal origin: chicory root, artichoke

  10. INULIN Biosensor advantages FOOD INDUSTRY Determination of interest in: inherent specificity simplicity • MONITORING OF PROCESSES • inuline extraction • fructose production rapidity real time analysis - QUALITY CONTROL - diethetic and children’s foods - component of dietary fiber • ECONOMIC AND LEGISLATIVE • added value for functional foods • ingredients establish prices

  11. 2TTF+ 2e PQQH2 5-CETO-D-FRUCTOSE 2TTF PQQ FRUCTOSE INULIN Redox mediator E = +0.2 V PBS 0.05 M, pH 4.5 BIENZYME BIOSENSOR FOR INULIN Gold nanoparticle preparation: By adding sodium citrate to a boiling HAuCl4 aqueous solution HAuCl4/sodium citrate  Particle size  Aucol TTF FDH AuE Cyst Inulinase

  12. 0.22 m i, A 0.20 0.18 0.16 +3s 0.14 0.12 -3s 0.10 0.08 0.06 0.04 0 20 40 60 80 100 120 140 160 time, days BIENZYME BIOSENSOR FOR INULIN Stability More than 5 months Storage conditions: 0.05 M phosphate buffer, pH 4.5, a 4ºC

  13. 20 nA 100 s BIENZYME BIOSENSOR FOR INULIN Interferences dextrose glucose lactose maltose saccharose inulin additions 10% Erel [INTERFERENT] / [INULIN] = 1

  14. 15 i, nA 12 9 6 3 0 0 2 4 6 8 10 12 14 16 18 20 t, min Chicory powder (18.5% inulin) 19.5 ± 0.4%, RSD = 2%, n = 6 Prebiotic food “Mas Vital”(2.0% inulin) 1.8 ± 0.1%, RSD = 5%, n = 6 BIENZYME BIOSENSOR FOR INULIN Sample preparation by size exclusion SPE Sample inulin Bio-Gel P-6 (Bio-Rad) fructose 2 μA t, min

  15. m A i, 16 12 8 4 0 -2 0 200 400 600 800 1000 E, mV COLLOIDAL GOLD-CARBON NANOTUBES COMPOSITE ELECTROCHEMICAL BIOSENSORS Hybrid nanoparticles/nanotubes materials Biocompatible materials with important electroanalytical features Aucoll-CNT-Teflon electrode graphite-Teflon (30:70) CNTs-Teflon (30:70) Aucoll-CNTs-Teflon Slope values of the calibration plot over (1.0-5.0)x10-3 M H2O2, at Eapp=+0.5 V 0.0083μA mM-1; 2.1 μA mM-1; 4.3 μA mM-1 Other advantages: Much lower noise level Rapidity J. Electroanal. Chem., 603 (2007) 1

  16. COLLOIDAL GOLD-CARBON NANOTUBES COMPOSITE ELECTROCHEMICAL BIOSENSORS Analytical characteristics and kinetic parameters for glucose biosensors based on GOx–CNT electrodes BIOSENSOR Edet, V Linear range, Slope, LOD, Useful KMapp mM mA/M μM lifetime GOx-Aucoll-CNT-Teflon +0.5 vs Ag/AgCl 0.05 – 1 2.6 17 3 months 14..9 GOx-CNT-Teflon +0.5 vs Ag/AgCl 0.1-8 1.2 33 1day 30

  17. COLLOIDAL GOLD-CARBON NANOTUBES COMPOSITE ELECTROCHEMICAL BIOSENSORS Current, % 100 75 50 25 0 8 6 2 4 Days GOx –Aucoll –CNT-Teflon biosensor GOx –CNT-Teflon biosensor 1.0 x 10-3 M glucose; Eapp=+0.5 V

  18. Aucoll-CNT-Teflon Aucol CNT electrocatalytic activity Enhanced electrode kinetics Suitable electrode material for NADH detection Suitable for the preparation of dehydrogenase biosensors

  19. Aucoll-CNT-Teflon NADH amperometric detection enhanced currents at less positive potentials 50 40 30 20 10 i, µA 0 0.2 0.4 0.6 0.8 E, V Aucoll-CNT-Teflon Aucoll-graphite-Teflon CNT-Teflon graphite-Teflon

  20. repeatability RSD = 3.7 % (n=10) NADH, 2.0 x 10-4 M rapidity t90%= 10-15 s Aucoll-CNT-Teflon NADH amperometric detection 10 mA 50 s Aucoll-CNT-Teflon CNT-Teflon Aucoll-graphite -Teflon 0.5 mA 50 s graphite -Teflon 0.5 mA 50 s Eapp.=+0.3 V; NADH, 1.0 x 10-4M

  21. Analytical characteristics Aucoll-CNT-Teflon NADH DETECTION Redox mediator • the highest calibration plot slope value • low detection potential with no mediator

  22. Alcohol dehydrogenase biosensor based on a colloidal gold-carbon nanotubes composite electrode ADH-Aucoll-CNT-Teflon 2e NADH CH3CHO NAD+ ETHANOL CH3CH2OH ADH Electrochim. Acta, 53 (2008) 4007-4012

  23. ADH-Aucoll-CNT-Teflon Analytical characteristics ETHANOL DETERMINATION Redox mediator higher slope value even with no mediator

  24. ADH-AucolL-CNT-Teflon APPLICATION sample RESULTS a)us stirring b)dilution CO2 Ethanol concentration, g / 100 ml* analytical solution Sample FREE Sample WITH Found Declared * mean value + ts / √n (n = 3)

  25. DETERMINATION OF GLUCOSINOLATE IN VEGETABLES Β-thioglucoside-N-hydroxysulfates Found in cabbage and broccoli Ingredient in functional foods Anticarcinogenic properties glucosinolates MYR H H O O 2 glucose H H O O 2e FAD FAD 2 2 2 GOx GOx O O FADH 2 2 2 MYR/GOx-Aucoll-CNT-Teflon Electroanalysis, 21 (2009) 1527

  26. CONCLUSIONS Gold nanoparticles allows the construction of electrochemical biosensors exhibiting enhanced performances with respect to other designs The unique properties of gold nanoparticles concerning immobilization of biomolecules retaining their biological activity, and as efficient conducting interfaces with electrocatalytic ability makes them a powerful tool to modify electrode materials and to construct robust and sensitive biosensors. They can be powerful analytical tools to be applied to the food industry. Applications in this field comprise the whole food chain, from the primary production to the final distribution to the consumer, which implies an enormous potential of application to food traceability.

  27. Thank you

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