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Seo and Shin: Quantitative Analysis of Eleven Bioactive Constituents of a Traditional Herbal Medicine, Yeonggyechulgam-tang using, Liquid Chromatography-Electrospray Ionization Tandem Mass Spectrometry
Original Article

Natural Product Sciences 2017;23(2):84-91.

Published online: 20 January 2017

DOI: https://doi.org/10.20307/nps.2017.23.2.84

Quantitative Analysis of Eleven Bioactive Constituents of a Traditional Herbal Medicine, Yeonggyechulgam-tang using, Liquid Chromatography-Electrospray Ionization Tandem Mass Spectrometry

Chang-Seob Seo, Hyeun-Kyoo Shin∗

K-herb Research Center, Korea Institute of Oriental Medicine, 1672 Yuseong-daero, Yuseong-gu, Daejeon 34054, Korea

∗Author for correspondence Dr. Hyeun-Kyoo Shin, K-herb Research Center, Korea Institute of Oriental Medicine, 1672 Yuseong-daero, Yuseong-gu, Daejeon 34054, Korea. Tel: +82-42-868-9464; E-mail: hkshin@kiom.re.kr

Received 25 November 2016       Revised 19 January 2017       Accepted 23 January 2017

Copyright © 2017 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

This study proposes a sensitive and selective liquid chromatography-electrospray ionization tandem mass spectrometry method of efficiently assessing the quality of a traditional herbal medicine called Yeonggyechulgam-tang (YGCGT). The following compounds 1 – 11, namely, liquiritin apioside (1), liquiritin (2), liquiritigene (3), coumarin (4), cinnamic acid (5), cinnamaldehyde (6), glycyrrhizin (7), atractylenolide III (8), atractylenolide II (9), atractylenolide I (10), and pachymic acid (11) were separated on a UPLC BEH C18 column (2.1 × 100 mm, 1.7 μm) at a column temperature of 45oC eluted with a gradient condition of 0.1% (v/v) formic acid in distilled water and acetonitrile. The correlation coefficient of the calibration curve of the eleven constituents was ≥ 0.9936. The limits of detection and quantification of the compounds 1 – 11 were 0.06 – 4.73 ng/mL and 0.17–14.20 ng/ mL, respectively. Using this analytical method, the compound 11 in lyophilized YGCGT decoction extract was not detected, while the compounds 1 – 10 were detected 0.13–166.43 mg/g.

Keywords: Quantitative analysis, Yeonggyechulgam-tang, LC-MS/MS

References

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Fig. 1.
Chemical structures of the compounds 1 – 11 in Yeonggyechulgam-tang.
nps-23-84f1.tif
Fig. 2.
Mass spectra of the precursor ion (Q1, A) and product ion (Q3, B) for LC-MS/MS MRM mode of the compounds 1 – 11. Liquiritin apioside (1), liquiritin (2), liquiritigene (3), coumarin (4), cinnamic acid (5), cinnamaldehyde (6), glycyrrhizin (7), atractylenolide III (8), atractylenolide II (9), atractylenolide I (10), and pachymic acid (11).
nps-23-84f2.tif
Fig. 3.
Total ion chromatograms of the reference standard (A) and lyophilized Yeonggyechulgam-tang extract (B) by LC-MS/MS MRM mode. Liquiritin apioside (1), liquiritin (2), liquiritigene (3), coumarin (4), cinnamic acid (5), cinnamaldehyde (6), glycyrrhizin (7), atractylenolide III (8), atractylenolide II (9), atractylenolide I (10), and pachymic acid (11, not detected).
nps-23-84f3.tif
Table 1.
Composition of Yeonggyechulgam-tang
Herbal medicine Scientific name Origin Amount (g)
  Poria Sclerotium Poria cocos Wolf Pyeongchang, Korea 7.500
  Cinnamomi Ramulus Cinnamomum cassia Presl Vietnam 5.625
  Atractylodis Rhizoma Alba Atractylodes macrocephala Koidzumi China 5.625
  Glycyrrhizae Radix et Rhizoma Glycyrrhiza uralensis Fischer China 3.750
Total amount     22.500
Table 2.
Linearities, regression equation, correlation coefficients, LOD, and LOQ for the compounds 1 – 11
Analyte Linear range (ng/mL) Regression equationa Correlation coefficient LODb (ng/mL) LOQc (ng/mL)
1 10–500 y = 8.21x – 27.85 0.9997 0.55 1.65
2 10–500 y = 9.01x – 19.10 1.0000 1.14 3.41
3 10–500 y = 29.58x – 29.21 1.0000 0.11 0.34
4 10–500 y = 38.59x + 51.41 0.9999 0.91 2.74
5 10–500 y = 21.35x – 67.24 0.9999 0.87 2.60
6 10–500 y = 1.15x – 18.20 0.9936 4.73 14.20
7 10–500 y = 3.08x – 60.81 0.9961 1.42 4.27
8 10–500 y = 52.12x – 97.00 1.0000 0.15 0.44
9 10–500 y = 56.54x + 33.78 1.0000 0.06 0.17
10 10–500 y = 53.42x + 63.53 1.0000 0.12 0.37
11 10–500 y = 1.46x – 10.67 1.0000 1.05 3.16

a y: peak area of compounds; x: concentration (ng/mL) of compounds.

b LOD = 3 × signal-to-noise ratio.

c LOQ = 10 × signal-to-noise ratio.

Table 3.
Mass detection condition of the compounds 1 – 11
Analyte Molecular weight (Da) Ionization mode Retention time (min) Precursor ion (m/z) Product ion (m/z) Cone voltage (V) Collision energy (eV)
1 550.5 [M−H]− 1.53 549.3 255.0 45 30
2 418.4 [M−H]− 1.65 417.4 255.2 30 15
3 256.3 [M+H]+ 2.99 257.2 137.0 35 25
4 146.1 [M+H]+ 3.06 147.1 91.0 30 20
5 148.1 [M+H]+ 3.74 148.9 130.9 20 20
6 132.1 [M+H]+ 4.42 133.1 115.0 25 15
7 822.9 [M−H]− 5.25 821.9 351.2 45 40
8 248.3 [M+H]+ 6.73 249.3 231.2 25 10
9 248.3 [M+H]+ 8.26 233.2 187.1 35 15
10 230.3 [M+H]+ 9.34 231.2 185.1 35 20
11 528.8 [M−H]− 11.97 527.6 465.4 45 35
Table 4.
Amounts of the compounds 1 – 11 in lyophilized Yeonggyechulgam-tang extract (n = 3)
Compound Amount (mg/g) Source
Mean SD RSD (%)
1 81.58 1.28 1.56 G. uralensis
2 79.88 0.31 0.39 G. uralensis
3 8.96 0.27 3.02 G. uralensis
4 54.52 1.45 2.65 C. cassia
5 10.18 0.25 2.50 C. cassia
6 121.09 1.11 0.92 C. cassia
7 166.43 11.17 6.71 G. uralensis
8 2.47 0.04 1.58 A. macrocephala
9 0.88 0.03 2.94 A. macrocephala
10 0.13 0.01 4.44 A. macrocephala
11 N.D.a – – P. cocos

a N.D. means not detected.

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