180
G. Saha et al.
2.4 Fatty Acid Analysis of Esterified Phospholipid and Oil
Isolated products were methylated by adding 1 mL diethyl ether and 1 mL of 0.5 N
methanolic KOH, followed by shaking the mixture for 10 min vigorously. HCl (1 mL,
1 N) was added, and the methyl esters of fatty acids were extracted with petroleum
ether (40–60 °C) (Litchfield 1972). Gas chromatographic analysis was next done as
follows.
2.5 Gas Chromatography
Fatty acid methyl esters were analyzed on a Hewlett-Packard gas chromatograph
(HP 5890A) (7890B GC system of Agilent Technologies), equipped with a flame
ionization detector (FID) and capillary DB-Wax column (30 mL, 0.250 mm I.D,
0.25 µm FT). The column temperature was programmed between 150 and 240 °C.
The different fatty acid methyl esters and the standard sample were separated on the
same column under identical conditions. N 2 as a carrier gas was used at 25 mL/min
(Saha et al. 2018).
2.6 Antioxidant Activity Determination by 1,
1-Diphenyl-2-Picrylhydrazyl (DPPH) Radical Inhibition
Assay
DPPH assay was determined by method of Blois (1958) and modified by Saha et al.
(2018). Each sample was added to 1 mL of 1 mM DPPH in chloroform. The decrease
in absorbance was monitored at 517 nm until a constant reading was obtained. The
readings were compared with the controls, which contained solvent (chloroform)
instead of the extract. The percentage inhibition was calculated as follows:
% inhibition =
Absorbance of control − Absorbance of sample
Absorbance of control
× 100
2.7 Total Phenolic Assay
The total phenolic content (TPC) was determined by the assay method modified
by Yu et al. (2003) and then also modified by Saha et al. (2018). Briefly, 1 ml of
solvent extract was transferred into a test tube and mixed with 1 mL of 95% ethanol
and 5 mL of dH 2 O. Folin–Ciocalteu reagent (50% v/v; 0.5 mL) was added to each
G. Saha et al.
2.4 Fatty Acid Analysis of Esterified Phospholipid and Oil
Isolated products were methylated by adding 1 mL diethyl ether and 1 mL of 0.5 N
methanolic KOH, followed by shaking the mixture for 10 min vigorously. HCl (1 mL,
1 N) was added, and the methyl esters of fatty acids were extracted with petroleum
ether (40–60 °C) (Litchfield 1972). Gas chromatographic analysis was next done as
follows.
2.5 Gas Chromatography
Fatty acid methyl esters were analyzed on a Hewlett-Packard gas chromatograph
(HP 5890A) (7890B GC system of Agilent Technologies), equipped with a flame
ionization detector (FID) and capillary DB-Wax column (30 mL, 0.250 mm I.D,
0.25 µm FT). The column temperature was programmed between 150 and 240 °C.
The different fatty acid methyl esters and the standard sample were separated on the
same column under identical conditions. N 2 as a carrier gas was used at 25 mL/min
(Saha et al. 2018).
2.6 Antioxidant Activity Determination by 1,
1-Diphenyl-2-Picrylhydrazyl (DPPH) Radical Inhibition
Assay
DPPH assay was determined by method of Blois (1958) and modified by Saha et al.
(2018). Each sample was added to 1 mL of 1 mM DPPH in chloroform. The decrease
in absorbance was monitored at 517 nm until a constant reading was obtained. The
readings were compared with the controls, which contained solvent (chloroform)
instead of the extract. The percentage inhibition was calculated as follows:
% inhibition =
Absorbance of control − Absorbance of sample
Absorbance of control
× 100
2.7 Total Phenolic Assay
The total phenolic content (TPC) was determined by the assay method modified
by Yu et al. (2003) and then also modified by Saha et al. (2018). Briefly, 1 ml of
solvent extract was transferred into a test tube and mixed with 1 mL of 95% ethanol
and 5 mL of dH 2 O. Folin–Ciocalteu reagent (50% v/v; 0.5 mL) was added to each