Antioxidant Activity

Determination of antioxidant activity

Scavenging DPPH radicals

DPPH (2,2-diphenyl-1-picryl-hydrazyl-hydrate) radical is used free radical method is an in antioxidant assay forwas used to evaluate measured the free radical scavenging activity of the lichen extract [8]. Two millilitreers of 0 .05 mg/mL methanol solution of DPPH radical in the concentration of (0 .05 mg/mL) and 1 mL of the lichen extract (1 mg/mL) were placed in cuvettes. The mixture is storewas stored stand at room temperature for 30 min. Then, the absorbance was measured at 517 nm in a spectrophotometer (Jenway, UK). Ascorbic acid was used as a positive control. DPPH radical activity (D) was calculated using the following equation:

Dt (%) = [(A0 - A1)/A0]x}100 where Ao

One millilitreer of the lichen extract (1 mg/mL) in a volumetric flask was diluted with distilled water (46 mL), and the content was mixed in a volumetric flask after adding. oOne millilitreer of Folin-Ciocalteu reagent was added and the content of the flask was mixed. thoroughly . After 3 min, 3 mL of 2% sodium carbonate (2 %) was added and then was allowed to standleft for 2 h with intermittent shaking. The reaction absorbance of prepared mixture absorbance was measured at 760 nm in a spectrophotometer (Jenway UK). The total concentration of phenolic compounds in the extract was determined expressed as microgram of pyrocatechol equivalent (PE) per milligram of driedy extract. The total phenolics compoundscontent was determined as the pyrocatechol equivalent using an equation obtained from a standard pyrocatechol graph (y = 0.0057 x total phenols [μg PE/mg of dry extracts] - 0.1646,

for the determination of the minimum inhibitory concentration (MIC) of the active extract. The sStarting solutions of the tested extract were obtained by dissolving it in 5% dimethyl-sulphoxide. STwo fold serial dilutions twofold dilutions of the extract were made within a concentration range from 0.04 to 40 mg/mL in sterile 96-well plates containing Mueller–Hinton broth for bacterial cultures and a Sabouraud Dextrose SD broth for fungal cultures. Resazurin solution was added as an indicator to each well. and finally, to each well fungal or bacterial suspension was added . The MIC was determined visually and defined as the lowest concentration of the tested extract that prevented resazurin colour change from blue to pink. Strep¬tomycin and ketoconazole were used as positive controls, while 5% DMSO was used as the a negative