Induction, Multiplication, and Evaluation of Antioxidant Activity of Polyalthia bullata Callus, a Woody Medicinal Plant

Abstract

Polyalthia bullata is an endangered medicinal plant species. Hence, establishment of P. bullata callus culture is hoped to assist in mass production of secondary metabolites. Leaf and midrib were explants for callus induction. Both of them were cultured on Murashige and Skoog (MS) and Woody Plant Medium (WPM) containing different types and concentrations of auxins (2,4-dichlorophenoxyacetic acid (2,4-D), α-naphthaleneacetic acid (NAA), picloram, and dicamba). The callus produced was further multiplied on MS and WPM supplemented with different concentrations of 2,4-D, NAA, picloram, dicamba, indole-3-acetic acid (IAA), and indole-3-butyric acid (IBA) media. The quantification of total phenolic content (TPC), total flavonoid content (TFC) and antioxidant capacity was further carried out on P. bullata callus, and the results were subjected to correlation analysis. Among the media, the WPM + 16.56 µM picloram (53.33 ± 22.06%) was the best for callus induction while MS + 30 µM dicamba was the best for callus multiplication. The TPC, TFC, and EC₅₀ of DPPH scavenging activity were determined at 0.657 ± 0.07 mg GAE/g FW, 0.491 ± 0.03 mg QE/g, and 85.59 ± 6.09 µg/mL in P. bullata callus, respectively. The positive correlation between DPPH scavenging activity with TPC was determined at r = 0.869, and that of TFC was at r = 0.904. Hence, the P. bullata callus has an ability to accumulate antioxidants. It therefore can be a medium for secondary metabolites production.

Keywords: Polyalthia bullata; antioxidant activity; auxins; callus induction; callus multiplication; total flavonoid content; total phenolic content.

Figure 1

Fresh weight and dry weight of P. bullata callus grown on MS medium at different concentrations of (a,b) α-naphthaleneacetic acid (NAA), (c,d) indole-3-butyric acid (IBA), (e,f) indole-3-acetic acid (IAA), (g,h) 2,4-D, (i,j) picloram, and (k,l) dicamba. Data represented as means ± SE. The asterisk (*) represents a significant difference of treated callus with 1-week old callus grown on MS basal media without auxins (MSO) at p ≤ 0.05 Tukey’s range test.

Figure 1

Fresh weight and dry weight of P. bullata callus grown on MS medium at different concentrations of (a,b) α-naphthaleneacetic acid (NAA), (c,d) indole-3-butyric acid (IBA), (e,f) indole-3-acetic acid (IAA), (g,h) 2,4-D, (i,j) picloram, and (k,l) dicamba. Data represented as means ± SE. The asterisk (*) represents a significant difference of treated callus with 1-week old callus grown on MS basal media without auxins (MSO) at p ≤ 0.05 Tukey’s range test.

Figure 2

Fresh weight and dry weight of P. bullata callus grown on WPM medium at different concentrations of (a,b) NAA, (c,d) IBA, (e,f) IAA, (g,h) 2,4-D, (i,j) picloram and (k,l) dicamba. Data represented as means ± SE. The asterisk (*) represents a significant difference of treated callus with 1-week old callus grown on WPM basal media without auxins (WPMO) at p ≤ 0.05 Tukey’s range test.

Figure 2

Fresh weight and dry weight of P. bullata callus grown on WPM medium at different concentrations of (a,b) NAA, (c,d) IBA, (e,f) IAA, (g,h) 2,4-D, (i,j) picloram and (k,l) dicamba. Data represented as means ± SE. The asterisk (*) represents a significant difference of treated callus with 1-week old callus grown on WPM basal media without auxins (WPMO) at p ≤ 0.05 Tukey’s range test.

Figure 3

Morphology of callus cultured on MS and WPM media supplemented with (a) MSO, (b) MS + 40 µM NAA, (c) MS + 50 µM IBA, (d) MS + 20 µM IAA, (e) MS + 30 µM 2,4-D, (f) MS+ 20 µM picloram, (g) MS + 30 µM dicamba, (h) WPMO, (i) WPM + 20 µM NAA, (j) WPM + 40 µM IBA, (k) WPM + 20 µM IAA, (l) WPM + 40 µM 2,4-D, (m) WPM + 10 µM picloram, and (n) WPM + 40 µM dicamba after six weeks of cultivation in dark condition at 25 ± 2 °C.

Figure 4

The 2,2′-diphenyl-1-picrylhydrazyl (DPPH) scavenging activity at different concentrations of ascorbic acid and methanolic extract of P. bullata callus.

Figure 5

The EC₅₀ value of DPPH scavenging activity of ascorbic acid and methanolic extract of P. bullata callus. The results expressed as the means ± standard error (n = 3). Means with different letters indicate significant difference at p ≤ 0.05.