In Vitro Antibacterial And Wound Healing Properties Of A Standardized Polyphenols-Rich Fraction Of Dicranopteris Linearis (BURM.) Underwood
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Date
2016-03
Authors
Ponnusamy, Yasodha
Journal Title
Journal ISSN
Volume Title
Publisher
Universiti Sains Malaysia
Abstract
Dicranopteris linearis (Burm.)Underwood is a ground creeping fern used
traditionally to treat various skin related ailments, such as boils, ulcers and external
wounds. To date, no scientific studies have been carried out to evaluate its
ethnopharmacological use in wound infection in a systematic manner. In the present
study, the MeOH extract of D. linearis was fractionated with the bioactive
constituents following a bioactivity-guided approach. A fraction (F5) of the extract
was obtained and it was found to possess significant antioxidant and antibacterial
activity against S. aureus and P. aeruginosa. Analysis was further carried out to
identify the chemical constituents of this active fraction. Isolation of the major
constituents was carried out using a combination of reversed-phase and normal-phase
silica gel chromatographic techniques. Two phenolic compounds which are
taxonomically significant to D. linearis, namely dichotomain A (2) and dichotomain
B (3), and two new compounds, 4-vinyl phenol-1-O-(2‟-O-α-L-rhamnopyranosyl)-β-
D-glucopyranoside (5) and 4-vinyl phenol-1-O-(4‟-O-α-L-rhamnopyranosyl)-β-Dglucopyranoside
(6) together with two other known compounds kaempferol 3-O-β-
D-glucopyranoside (1) and kaempferol 3-O-β-D-glucosyl (1→2)-β-glucoside (4)
were obtained. Characterization of the constituents was carried out using various
spectroscopic techniques such as UV, IR, NMR and MS. In order to determine the
content of F5 and to standardize the fraction, a novel HPLC–DAD method was developed and validated using the isolated compounds as analytical markers. This
method was found to be accurate and precise, with an LOQ of 1.56 μg/mL for
compounds 1, 4, 5& 6 and 15.6 μg/mL for compound 2. The contents of the marker
compounds were found to be 3.1% for compound 1, 3.4% for compound 2, 3.4 % for
compound 4, and 26.8 % for compound 5 & 6. Antibacterial and wound healing
potential of the standardized F5 fraction of D. linearis was subsequently investigated
using several in vitro assays. From the time killing assay, minimum inhibitory
concentration (MIC) assay and scanning electron microscopic images, F5 was found
to exert bacteriostatic effect against S. aureus and P. aeruginosa. Moreover, F5 was
also able to potentiate the antibacterial effect of few conventional antibiotics,
namely, chloramphenicol, penicillin G and ampicillin against S. aureus and MRSA.
On the other hand, F5 was able to enhance the proliferation of fibroblast cells and
induce cell migration in the scratch-wound assay. In addition, it protected the cells
against hydrogen peroxide induced oxidative stress and was non-cytotoxic towards
both human and mouse fibroblast cell lines up to the tested concentrations of
500μg/mL. Taken together, the present study provided novel information to support
the traditional use of this plant for skin infections. In vivo studies are warranted to
further explore the potential of D. linearis as a wound healing agent.
Description
Keywords
Antioxidant and antibacterial