3.2.2. Antioxidant Activities

The antioxidant activities of *B. glabra* nodal extracts, propagated via conventional method and in vitro-induced calli via tissue culture techniques, were measured using three in vitro antioxidant assays including 2,2-diphenyl-1-picryl-hydrazyl-hydrate (DPPH) free radical scavenging activity, 2,2-azino-bis (3-ethylbenzothiazoline-6-sulfonic acid) (ABTS), and iron (II) chelating activity. All antioxidant assays were measured using a UV-Vis spectrophotometer at a specific absorbance. The results showed that DPPH free radical scavenging activity and ABTS scavenging activity of the conventionally propagated nodal segment were significantly different at *p* ≤ 0.05 than in vitro-induced calli under dark and light incubation condition. However, the highest iron (II) chelating activity was obtained from in vitro-induced calli under a dark incubation condition. The DPPH free radical scavenging activity recorded from sources of samples and extraction solvents ranged between 0.14 to 7.64 mg TE/g DW. The highest DPPH free radical scavenging activity was obtained from aqueous extract of the node, followed by ethanol extract of the node, and aqueous extract of the calli induced under photoperiod. In addition, there were no significant differences between hexane extract with all of the samples. The lowest DPPH free radical scavenging activity was recorded from hexane extract of calli induced under photoperiod and dark conditions.


**Table 4.** 2,2-diphenyl-1-picryl-hydrazyl-hydrate (DPPH) free radical scavenging activity and 2,2 azino- bis (3-ethylbenzothiazoline-6-sulfonic acid) (ABTS) scavenging activity of node and in vitroinduced calli of *Bougainvillea glabra*.

Values are means ± SE, the same letter within the same column for each factor indicates no significant difference (*p* < 0.05). The means separation is done using Duncan's multiple range test (DMRT).



Values are means ± SE, the same letter within the same column for each factor indicates no significant difference (*p* < 0.05). The means separation is done using Duncan's multiple range test (DMRT).

In the analysis of ABTS scavenging activity, there was a significant interaction between the sources of samples and extraction solvents on ABTS scavenging activity. The highest ABTS scavenging activity was significantly obtained by aqueous extract of the node followed by aqueous extract of the calli induced under the photoperiod, and aqueous extract of the calli induced under the dark condition Meanwhile, there were no significant differences between ethanol extract of the node and calli induced under photoperiod conditions. In addition, there were no significant differences between ethanol, acetone, and hexane extract of the calli induced under dark conditions. On the other hand, the lowest ABTS scavenging activity was recorded from hexane extract of calli induced under dark conditions. Thus, all of these extracts had some ABTS radical scavenging abilities in their phytochemical components, and the contents may have some ABTS radical scavenging ability equivalence.

In the present study on iron (II) chelating activity, a significant interaction was found between extraction solvents and sources of samples. In contrast to other antioxidant activities conducted in which aqueous and other node extracts produced higher antioxidant activities, the iron (II) chelating activity was produced from the aqueous extract of the calli induced under dark conditions. The highest iron (II) chelating activity was produced from an aqueous extract of the calli induced under the dark condition. Meanwhile, there were no significant differences between the aqueous extract of the node and calli induced under photoperiod and hexane extract of the calli induced under dark conditions. In addition, there were no significant differences between ethanol extract of the node, calli induced under photoperiod, and dark conditions. Furthermore, the lowest iron (II) chelating activity was produced by the acetone extract of calli induced under the photoperiod

#### 3.2.3. Correlation Analysis between Variables

Pearson's correlation analysis was used to examine the relationship between phenolic contents (phenolic acids and flavonoids) and antioxidant activities, such as DPPH free radical scavenging activity, ABTS scavenging activity, and iron (II) chelating activity (Table 6).


**Table 6.** Pearson's correlation analysis between variables.

Notes: \*\*: Significant correlation at *p* < 0.05; ns: Non-significant correlation; TPC: Total phenolic acids content; TFC: Total flavonoids content; DPPH: DPPH free radical scavenging activity; ABTS: ABTS scavenging activity; Fe2+: Iron (II) chelating activity, respectively.

The correlation showed that all variables except iron (II) chelating activity were positively correlated ranged from intermediate positive to a high positive correlation. The variables of total phenolic acids, flavonoids, DPPH free radical scavenging activity, and ABTS scavenging activity produced a highly significant strong correlation (r > 0.75). Meanwhile, iron (II) chelating activity showed a low positive correlation against TPC and ABTS (r < 0.75) and a non-significant correlation against TFC and DPPH free radical scavenging activity. The Pearson's correlation analysis showed that the phenolic compounds present in *B. glabra* nodes and calli induced under different light condition extract are strong as scavenging and chelating agents.

## **4. Discussion**
