Composition and Antioxidant Activity of Vapours …
153
2.2 Determination of Total Polyphenols (TPC)
Total polyphenols content (TPC) was determined by Singleton et al. (1999) method.
2.3 Determination of Antioxidant Activity
2.3.1 DPPH Free Radical Scavenging Activity Assay
The total antioxidant capacity of two samples was determined spectrophotometrically, assessed using 1, 1-diphenyl 2-picryl-hydrazyl (DPPH) as followed by Shimada
et al 1992. 0.1 m mol solution of DPPH in methanol was prepared. The absorbance
was measured 30 min after the initial mixing of samples with 1 ml of DPPH solution at 517 nm. The ability to scavenge the DPPH radical was calculated with the
following equation:
Inhibition percentage (I%) = (A 0 − A 1 )/A 0
(A 0 = absorbance of the control, A 1 = absorbance of the sample).
2.3.2 Ferric Reducing Antioxidant Power (FRAP)
The ferric ions (Fe
3+ ) reducing antioxidant power (FRAP) method was used to
measure the reducing capacity of samples, described by Benzie and Strain 1996. The
FRAP reagent consists of 10 mM TPTZ in 40 mM HCL, 250 mM sodium acetate
buffer (pH-3.6) and 20 mM FeCl 3 . The reagent was freshly prepared by mixing TPTZ
solution, FeCl 3 solution and acetate buffer in a ratio of 1:1:10. Samples (100 μl) were
mixed with 900 μl of FRAP reagent.The mixture was incubated at 37 °C for 4 min
and the absorbance was measured at 593 nm.
2.3.3 ABTS Free Radical Scavenging Activity
ABTS
+ assays were done by modified procedure described by Re et al. 1999. ABTS
+
(ABTS radical cation) was generated through the reaction between potassium per
sulphate (2.45 μM) and ABTS stock solution. The mixture was kept in the dark place
for 16 h at room temperature before use. The resulting intense colour matches the
ABTS radical cations. The obtained solution was subsequently diluted with methanol
and absorbance was measured at 734 nm. 1 ml of ABTS diluted solution was mixed
with 10 μl of sample and the reaction mixture was kept for 6 min before measuring
the absorbance at 734 nm. ABTS scavenging activity was calculated by the following
equation:
153
2.2 Determination of Total Polyphenols (TPC)
Total polyphenols content (TPC) was determined by Singleton et al. (1999) method.
2.3 Determination of Antioxidant Activity
2.3.1 DPPH Free Radical Scavenging Activity Assay
The total antioxidant capacity of two samples was determined spectrophotometrically, assessed using 1, 1-diphenyl 2-picryl-hydrazyl (DPPH) as followed by Shimada
et al 1992. 0.1 m mol solution of DPPH in methanol was prepared. The absorbance
was measured 30 min after the initial mixing of samples with 1 ml of DPPH solution at 517 nm. The ability to scavenge the DPPH radical was calculated with the
following equation:
Inhibition percentage (I%) = (A 0 − A 1 )/A 0
(A 0 = absorbance of the control, A 1 = absorbance of the sample).
2.3.2 Ferric Reducing Antioxidant Power (FRAP)
The ferric ions (Fe
3+ ) reducing antioxidant power (FRAP) method was used to
measure the reducing capacity of samples, described by Benzie and Strain 1996. The
FRAP reagent consists of 10 mM TPTZ in 40 mM HCL, 250 mM sodium acetate
buffer (pH-3.6) and 20 mM FeCl 3 . The reagent was freshly prepared by mixing TPTZ
solution, FeCl 3 solution and acetate buffer in a ratio of 1:1:10. Samples (100 μl) were
mixed with 900 μl of FRAP reagent.The mixture was incubated at 37 °C for 4 min
and the absorbance was measured at 593 nm.
2.3.3 ABTS Free Radical Scavenging Activity
ABTS
+ assays were done by modified procedure described by Re et al. 1999. ABTS
+
(ABTS radical cation) was generated through the reaction between potassium per
sulphate (2.45 μM) and ABTS stock solution. The mixture was kept in the dark place
for 16 h at room temperature before use. The resulting intense colour matches the
ABTS radical cations. The obtained solution was subsequently diluted with methanol
and absorbance was measured at 734 nm. 1 ml of ABTS diluted solution was mixed
with 10 μl of sample and the reaction mixture was kept for 6 min before measuring
the absorbance at 734 nm. ABTS scavenging activity was calculated by the following
equation: