TOTAL POLYPHENOLIC CONTENT MEASUREMENT USING POLYPHENOL SENSORS

1. TOTAL POLYPHENOLIC CONTENT MEASUREMENT USING POLYPHENOL SENSORS GRUPO 19 Cristina Ruiz Martínez Sara Ugarte Cerrato

2. INDEX • Introduction • Mechanisms • Samples • Red wine • Olive oil • Other vegetable extracts • A new biosensor

3. Introduction • Mechanisms • Samples • Red wine • Olive oil • Other vegetable extracts • A new biosensor

4. Introduction POLYPHENOL COMPOUNDS malvidin quercetin • Natural antioxidants widely distributed in the plant kingdom • Important determination • physiological effects • employment as markers in taxonomic studies • their properties to food quality control

5. Introduction • Mechanisms • Samples • Red wine • Olive oil • Other vegetable extracts • A new biosensor

6. Mechanism • CLASSICAL METHODS • spectrophotometry • gas chromatography • liquid chromatography • capillary electrophoresis • Folin-Ciocalteau • RECENT METHODS • Biosensors: basedonenzymes • Advantages: • rapid response • cost-effectiveness • simplicity of operation and manufacturing, minimal sample pretreatment and solvent requirements.

7. Biosensors for polyphenols determination

8. Introduction • Mechanisms • Samples • Red wine • Olive oil • Other vegetable extracts • A new biosensor

9. SAMPLESRED WINE • Enzyme: laccase • Product of its oxidation: 1,2-benzoquinone reduced at the electrode. • Actual mechanism of reaction still unclear. • From spectroscopic and electron paramagnetic resonance (EPR) studies: 1º enzyme completely reduced 2º oxygen reduced to water

10. WINE SAMPLE • Immobilization in polyethersulfone membranes: • 1º Washings containing an excess of enzyme with acetate buffer. • 2º A quantity deposited on polyethersulfone membrane cut in the form of discs. • 3º Application to the electrode. • 4º The biosensor is dipped in the buffer. • 5º Injections of the sample or standard under magnetic stirring.

11. WINE SAMPLE • Conclusion: able to discriminate between catechin and caffeic acid but negligible responses when using with wine. • The complex matrix of the red wine samples interference in the response. • More research to overcome the deviations.

12. Introduction • Mechanisms • Samples • Red wine • Olive oil • Other vegetable extracts • A new biosensor

13. OLIVE OIL SAMPLE • Twosensors: • Biosensor based on the catalytic activity of the tyrosinase. Main advantages: -prior extraction pre-treatment analysis time not necessary eliminated decreased - Flow injection analysis - saving time - Semiautomization of - minimization of the entire procedure exposure to solvent vapors

14. OLIVE OIL SAMPLE Tyrosinase Pre-actived membrane Amperometric Gas Diffuse Electrode Gas permeable membrane Dialysis membrane Sensor Teflon O-ring

15. OLIVE OIL SAMPLE • Voltammetric sensor • Prior extraction • Using a disposable screen-printed sensor (SPE) • Reference compound: oleuropein • A calibration curve of oleuropein

16. OLIVE OIL SAMPLE • Electrode

17. Introduction • Mechanisms • Samples • Red wine • Olive oil • Other vegetable extracts • A new biosensor

18. VEGETABLE EXTRACTS • Enzyme: horseradish peroxidase. • Inmmobilization: silica-titanium. • Material with high chemical stability. • Improvement of the amperometric detection. • No significant influence of the matrix was observed.

19. VEGETABLE EXTRACTS • Mechanism: double displacement or ping-pong.

20. Introduction • Mechanisms • Samples • Red wine • Olive oil • Other vegetable extracts • A new biosensor

21. New biosensor • Based on laccase immobilized onto silver nanoparticles/multiwalled carbon nanotube/polyaniline gold electrode. • Immobilization on AgNPs/PANI/MWCNT/Au (gold) electrode through covalent coupling to construct an enzyme electrode for determination of polyphenols. • Employed for amperometric determination of total phenolic content in beverages and pharmaceutical formulation. • Conclusion: good biosensor, likely to overcome the problem of leakage of enzyme.

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