1. Comparison of manual and automatised TAC determination in fruits Anita Herndl Short Term Scientific Mission COST Action 863 9.10.-20.10.2006 University of Natural Resources and Applied Life Sciences, Vienna Institute of Applied Microbiology

2. STSM Program Methods: Extraction Methods TEAC: Manual Flow Injection Total Phenolic Content (Folin- Ciocalteu) Total Anthocyanin Content Host: Maurizio Battino Università Politecnica delle Marche, Ancona

3. TEAC Trolox Equivalent Antioxidant Capacity Scavenging of the colored ABTS radical cation (ABTS + ) ABTS + is generated chemically or enzymatically Antioxidants in the sample directly react with ABTS + forming ABTS (colorless) Decolorisation is measured at 734 nm at a fixed time point Results are expressed relative to TROLOX ABTS: 2,2'-azino-bis(3-ethylbenzthiazoline-6-sulphonic acid) ABTS

4. TEAC + Widely used Simple and fast No special equipment required Stability of ABTS + ABTS + is soluble in aqueous and organic solvents can be automated - ABTS + not naturally present in mammalians underestimation

5. FIA-TEAC The system consists of: HPLC pump Manual injection valve with a 5 µL loop Reaction coil UV/VIS Detector ABTS +: 4°C Reaction: 35°C Measurment: after 1,3 min Bompadre S., Leone L., Politi A., Battino M. Improved FIA-ABTS method for antioxidant capacity determination in different biological samples. Free Radic Res. 2004; 38(8):831-8

6. Total Phenol Content Folin-Ciocalteu-Reagent Molybdotungstonphosphate heteropolyanion Blue reaction products Measurment at 760 nm Gallic acid as reference standard Results expressed as mg Gallic acid equivalent/ 100 g FW

7. Total Anthocyanin Content Anthocyanin pigments change color with a change in pH Colored at pH 1 Colorless at pH 4,5 Sample equilibrated at different pH λ max : Determination by wavelength scan Standard: predominant anthocyanin in sample for strawberries: Pelargonidin

8. Results Variation coefficient [%] µmol TE/g FW apple 1 st Day1,227,46 2 nd Day1,816,76 3 rd Day4,887,20 Inter-Day5,04 7,10 1 st Day3,235,85 2 nd Day19,675,67 Inter-Day12,54 5,76 FIA-TEAC Manual TEAC Hydrophilic antioxidant capacity determined by manual TEAC and FIA-TEAC

9. Results Variation coefficient [%] µmol TE/g FW apple 1 st Day2,691,93 2 nd Day7,031,68 3 rd Day3,091,84 Inter-Day7,461,85 1 st Day4,891,09 2 nd Day22,910,48 Inter-Day43,370,79 FIA-TEAC Manual TEAC Lipophilic antioxidant capacity determined by manual TEAC and FIA-TEAC

10. Results Total Phenolic Content (Apple) Total Anthocyanin Content (Strawberry) Variation coefficient [%]mg GAE/100g FW Hydrophilic components5,90101,88 Lipophilic components10,5026,42 SampleVariation coefficient [%]mg Pg/100g FW 11,3335,80 23,1932,22

11. Concluding Remarks Training on analytical methods measuring the antioxidant capacity of fruits Hydrophilic and lipophilic components Comparison of a manual and an automated assay FIA-TEAC: relative parameters efficently controlled higher reproducibility

12. Thanks to COST Action 863 Chairman Bruno Mezzetti Maurizio Battino Sara Tulipani Margit Laimer

13. THANK YOU FOR YOUR ATTENTION