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Shin, Chung, Park, and Lee: Identification of Antioxidative Constituents from Polygonum aviculare using LC-MS Coupled with DPPH Assay
Original Article

Natural Product Sciences 2016;22(1):64-69.

Published online: 17 December 2016

DOI: https://doi.org/10.20307/nps.2016.22.1.64

Identification of Antioxidative Constituents from Polygonum aviculare using LC-MS Coupled with DPPH Assay

Hyeji Shin1, Hayeon Chung1, Byoungduck Park2, Ki Yong Lee1, ∗

1College of Pharmacy, Korea University, Sejong 339-700, Korea

2College of Pharmacy, Keimyung University, Daegu 704-701, Korea

∗Author for correspondence Ki Yong Lee, College of Pharmacy, Korea University, 2511 Sejong-ro, Sejong, Korea Tel: +82-44-860-1623; E-mail: kylee11@korea.ac.kr

Received 2 September 2015       Revised 25 September 2015       Accepted 29 September 2015

Copyright © 2016 The Korean Society of Pharmacognosy

This is an Open Access article distributed under the terms of the Creative Commons Attribution Non-Commercial License (http://creativecommons.org/licenses/by-nc/3.0) which permits unrestricted non-commercial use, distribution, and reproduction in any medium, provided the original work is properly cited.

Abstract

A method for simultaneously identifying antioxidative compounds was developed using time-based LC-MS coupled with DPPH assay regardless of the time consuming process. The methanolic extract of Polygonum aviculare (Polygonaceae) showed significant DPPH radical scavenging activity. Time-based DPPH assay for simultaneous identification of active compounds from the extracts of P. aviculare was used. Major peaks of ethyl acetate fraction of P. aviculare showed high DPPH radical scavenging activity. A simple phenolic compound (1) and six flavonoids (2-7) were isolated from the ethyl acetate fraction of P. aviculare by silica gel and sephadex LH-20 column chromatography. The structures of seven compounds were determined to be protocatechuic acid (1), catechin (2), myricitrin (3), epicatechin-3-O-gallate (4), avicularin (5), quercitrin (6), and juglanin (7) based on the analysis of the1H-NMR,13C-NMR and ESI-MS data. All compounds exhibited significant antioxidant activity on DPPH assay and active compounds were well correlated with predicted one.

Keywords: Polygonum aviculare, Flavonoids, LC-MS, DPPH assay

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Fig. 1.
HPLC chromatogram (254 nm) and time-dependent DPPH assay results of EtOAc fraction from P. aviculare (A), expansion of the HPLC chromatogram between 11–16 min (B), UV spectrum of each peaks (C).
nps-22-64f1.tif
Fig. 2.
Structures of the compounds isolated from P. aviculare.
nps-22-64f2.tif
Table 1.
DPPH radical scavenging activity of the extract and fractions from P. aviculare
  0.04 μ g/ml 0.2 μ g/ml 1.0 μ g/ml 5.0 μ g/ml 10.0 μ g/ml
total extract 10.0 ± 1.1 9.7 ± 1.7 29.8 ± 1.3 66.6 ± 2.1 89.0 ± 1.2
n-hexane fr. -0.8 ± 0.9 21.1 ± 1.8 56.2 ± 2.0 92.4 ± 2.3 94.8 ± 0.1
EtOAc fr. 38.1 ± 1.8 76.8 ± 0.5 93.8 ± 1.1 98 ± 0.16 96 ± 0.10
n-BuOH fr. 5.9 ± 0.3 39.7 ± 3.2 94.7 ± 0.2 95.1 ± 0.3 95 ± 0.4
water fr. 7.6 ± 2.5 1.2 ± 2.6 3.2 ± 4.3 9.1 ± 3.0 14.8 ± 1.2

Data represent the means ± S.D. from triplication.

Table 2.
LC-MS analysis of each peak in EtOAc fractions from P. aviculare
Peaks Retention time (min) λmax (nm) [M-H]− (m/z) Expected type Isolated compounds
1 5.429 260, 295 153.0255 phenolic acid compound 1
2 6.856 280 289.0714 catechin compound 2
3 12.423 260, 350 463.0882 flavonoid glycoside compound 3
4 12.649 275 441.0919 catechin gallate compound 4
5 13.503 260, 350 433.0786 flavonoid glycoside compound 5
6 13.676 260, 350 447.1020 flavonoid glycoside compound 6
7 14.216 260, 350 615.1096 galloyl flavonoid glycoside  
8 14.483 260, 350 417.0847 flavonoid glycoside compound 7
Table 3.
DPPH radical scavenging activity of isolated compounds from P. aviculare
Compounds IC50, μ Ma
1 19.91
2 15.19
3 10.22
4 13.31
5 10.33
6 13.98
7 89.91
Curcuminb 27.80

a Inhibitory concentration, 50%

b Positive control

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