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METHODS

In vitro chemical assays

Scavenging activity of antioxidant compounds (including plant extracts) against Reactive Oxygen Species (ROS) and Reactive Nitrogen Species (RNS):

 

  • Superoxide anion radical;

  • Hydrogen peroxide;

  • Hypochlorous acid;

  • Singlet oxygen;

  • Peroxyl radicals;

  • Nitric oxide;

  • Peroxynitrite.

Microplates used for the in vitro chemical assays

In vitro cellular assays

Isolated human neutrophils:

 

  • Viability studies: trypan blue and LDH assays;

  • Evaluation of apoptosis by morphology and flow cytometry (annexin-V/PI);

  • p53 determination;

  • Measurement of caspases activity;

  • Evaluation of the neutrophils' oxidative burst modulation;

  • Neutrophils' chemotaxis assay;

  • Inhibitory effect of the inflammatory enzyme 5-LOX;

  • Determination of the enzymatic type of inhibition of soybean LOX-1.

Isolated human neutrophils

Isolated human erythrocytes:

 

  • Lipid peroxidation (TBARS);

  • Hemoglobin oxidation;

  • Hemolysis;

  • Both forms of glutathione: reduced (GSH) and oxidized (GSSG).

 

Whole blood assays:

  • Cytokines determination;

  • Inhibitory effect of the inflammatory enzymes (COX-1 and COX-2).

Isolated human erythrocytes

(red portion)

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