In vitro callus induction and evaluation of antioxidant activity of Rhinacanthus nasutus (L.) Kurz

Abstract

Rhinacanthus nasutus (L.) Kurz is used in Thai traditional medicine for the treatment of skin diseases, ringworm, and eczema. This research studied the effects of cytokinin and auxins on callus induction and evaluated antioxidant activity of R. nasutus. Nodes, young, and mature leaf explants were cultured on MS medium containing 0, 1, 2, 3, and 4 mg/l kinetin (6-furfurylaminopurine) and 0, 1 mg/l 1-naphthaleneacetic acid (NAA), indole-3-acetic acid (IAA), and 2,4-dichlorophenoxyacetic acid (2,4-D) for 6 weeks to induce callus. Calli derived from nodes, young and mature leaves, and other plant parts were ultrasonically extracted with methanol to determine total phenolic content (TPC), total flavonoid content (TFC), and antioxidant activity by ferric reducing antioxidant power (FRAP), 2,2-diphenyl-1-picrylhtdrazyl (DPPH), and 2,2′-Azino-bis(3-ethylbenzthiazoline-6-sulfonic acid) (ABTS) assays. Nodal explants on MS medium containing 1 mg/l kinetin combined with 1 mg/l 2,4-D were most efficient in callus production with the average fresh and dry weight per explant of 2.29 ± 0.14 and 0.18 ± 0.01 g, respectively. Addition of kinetin combined with NAA or 2,4-D had a positive effect on callus induction from young and mature leaf explants. The leaf extract showed the highest TPC, TFC, FRAP, and IC50 of DPPH and ABTS assays (ca 113 mg GAE/g extract, 45 mg QE/g extract, 121 mg TE/g extract, 53 μg/ml and 14 μg/ml, respectively), followed by callus derived from nodes. Overall, phenolic content was higher than flavonoid content. A strong positive correlation was found between FRAP assay, TPC (r = 0.973), and TFC (r = 0.798), indicating that phenolic and flavonoid compounds are responsible for antioxidant activity of R. nasutus.