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equilibrated to 1.00 (±0.10) by diluting with deionized water. Free radical scavenging

activities were measured across a concentration gradient of the plant samples (6.25–100
•+
µg/ml) by mixing with the ABTS solution. After incubation for about 30 min at room
temperature on a shaker, absorbance was measured at 405 nm using UV-Visible
spectrophotometer (Evolution 201; Thermo scientific). ABTS •+ scavenging activity of

ascorbic acid (100 µM) was also run in parallel and compared with the plant samples. Free
radical scavenging activities were expressed in inhibition percent (IP) as follows:

IP= (A1-A2)/A1 X 100

•+
•+
where A1 is the absorbance of ABTS solution and A2 is the absorbance of ABTS solution
treated with samples/ascorbic acid. Concentrations of the samples required for 50%
scavenging of the free radical (IC50) were then determined by using GraphPad Prism 8

software.
DPPH Assay

1,1-diphenyl-2-picrylhydrazyl (DPPH) radical scavenging activity of the plant extracts
was performed by using methods as described by Dzoyem and Eloff (2015) [15]. As similar

to the previous method, different concentrations of the plant extracts (6.25–100 µg/ml) were
mixed properly with DPPH solution (0.1 mM; in methanol) and incubated for about 30 min at

room temperature in dark. Then the absorbance was measured at 517 nm using the UV–

•
Visible spectrophotometer. The DPPH solution was used as control and methanol was used
as blank. Subsequently, IP and IC50 values were determined as described above.

Phenolic contents
Phenolic contents in different parts of S. acmella were determined by Folin-Ciocalteu

method [16]. Initially, the plant extracts were mixed with 10% Folin-Ciocalteu reagent and
allowed to stand at room temperature for 5 min in dark. Then the solutions were mixed

properly with sodium carbonate (Na2CO3; 5% w/v) and incubated in dark for 2h. The

absorbance of the solutions was measured at 750 nm using UV-Vis spectrophotometer. Gallic
2
acid was used for constructing standard curve (Y=0.009X; R =0.998) and the concentration
of phenolics in the plant samples was expressed as milligrams of Gallic acid equivalent per

gram (mg GAE/g) of dry extract.
Flavonoid contents

Flavonoid content in the plant extracts was determined by using aluminium chloride
colourimetric methods [17]. The plant extracts were mixed with sodium nitrite (NaNO2; 5%

w/v) and allowed to stand for 6 min. Then the solutions were mixed with aluminium
trichloride (AlCl3, 10% w/v) and allowed to stand for another 6 minutes. Later sodium

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