1. DPPH FREE RADICAL SCAVENGING ACTIVITY ANALYSIS BY SPECTROPHOTOMETRIC METHOD Dr. Nur CEBI

2. Antioxidant Activity of Foods  The antioxidant properties of each food matrix come from the combined and concertedaction of biologically active compounds, i.e., polyphenols, carotenoids, lignans, glucosinolates, etc.  Antioxidants can exert large spectra of biological and physiological functions, i.e., anti-allergic,  Anti-atherogenic, anti-inflammatory, antimicrobial, antioxidant, anti-thrombotic, cardioprotective, etc. and assessment of interactions between these biologically active compounds and other food matrix  Components could be seen as the main step in the investigation of total antioxidant properties, in terms of both the potential health benefits of food and as the indicator of a “possible beneficial role”.

3.  Antioxidants are found in many foods such as fruits and green leafy vegetables, and various forms of polyphenols or phytochemicals such as tannins, resveratrol, flavonoids, ellagic acid, catchechin etc.

6. Measurement of Antioxidant Activity of Foods  Oxygen radical absorbance capacity (ORAC assay)  The total radical-trapping antioxidant parameter (TRAP assay)  Ferric reducing antioxidant power (FRAP assay)  Cupric ions (Cu2) reducing antioxidant power(CUPRAC assay)  DPPH scavenging assay

7. DPPH scavenging assay  Aim: The antioxidant activity of the plant tor fruit extracts and the standard is evaluated on the basis of the radical scavenging effect by using a spectrophotometric procedure.  Principle: The ability of the extracts to annihilate the DPPH radical (1,1-diphenil- 2-picrylhydrazyl) was investigated.

8. DPPH scavenging assay  Antioxidants react with DPPH, which is a stable free radical and is reduced to the DPPH and as consequence there is decreased from the DPPH radical to the DPPH-H form.  The degree of discoloration indicates the scavenging potential of the antioxidant compounds or extracts in terms of hydrogen donating ability

9. DPPH scavenging assay  The scavenging reaction between (DPPH) and an antioxidant (H-A) can be written as:

12. DPPH scavenging assay  Chemicals:  DPPH (2,2-diphenyl-1-picryl-hydrazyl- hydrate) Reagent  Methanol (Analytical Grade)  UV-visible spectrophometer  Micropipette

13. DPPH scavenging assay  Experimental Procedure:  0.1mM of DPPH was prepared in methanol and 4.9 mL of DPPH (0.1mM) solution was mixed with 0.1 mL of sample solution (fruit or plant extract) and standard solution in test tubes separately in triplicates. These solution mixtures shanked vigorously, then were allowed to stand at dark for 30 min and optical density was measured at 517 nm using UV-VIS Spectrophotometer. Methanol (0.1 mL) with DPPH solution (0.1mM, 4.9 mL) was used as blank.

14. DPPH scavenging assay  DPPH Reagent: Methanolic solution of DPPH (0.1 mM): 39.4 mg of DPPH was dissolved (Ma=394,32 g/mol) m=39,432 g in one liter of analytical grade methanol 0.1 mM, 0.05 mM, 0.025 mM, 0.0125 mM M1xV1=M2xV2

15. DPPH solution concentration (mM)Absorbance Value (517 nm) 0,01250,121 0,0250,245 0,050,477 0,10,956 SamplesAbsorbance Values control0,947 Extractf0,864 Extract s0,399

16. Results

17. Report SamplesAbsorbance Values control0,947 Extract10,864 Extract 20,399 Extract30,225 kırmızı lahana eşleştir? patates kereviz

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