- Email: [email protected]
Rapid characterization of the chemical constituents of Sijunzi decoction by UHPLC coupled with Fourier transform ion cyclotron resonance mass spectrometry
Journal of Chromatography B 1086 (2018) 11–22
Contents lists available at ScienceDirect
Journal of Chromatography B journal homepage: www.elsevier.com/locate/jchromb
Review
Rapid characterization of the chemical constituents of Sijunzi decoction by UHPLC coupled with Fourier transform ion cyclotron resonance mass spectrometry ⁎
Zhibo Guana, Miao Wangb, Yi Caia, Hongmei Yanga, Min Zhaoa, , Chunjie Zhaoa, a b
T
⁎
School of Pharmacy, Shenyang Pharmaceutical University, Wenhua Road 103, Shenyang, Liaoning Province, China School of Life Science and Biopharmaceutics, Shenyang Pharmaceutical University, Wenhua Road 103, Shenyang, Liaoning Province, China
A R T I C LE I N FO
A B S T R A C T
Keywords: Sijunzi decoction Poria Atractylodis Macrocephalae Rhizoma Fourier transform ion cyclotron resonance Chemical constituents
Sijunzi decoction, a renowned Chinese prescription has long been utilized to treat gastrointestinal problems. In the context of this research work, the use of Ultra high performance liquid chromatography combined with Fourier transform ion cyclotron resonance mass spectrometry was made to separate and characterize the components of Sijunzi decoction. The performance of Liquid chromatography was carried out on a C8 column (150 mm × 2.1 mm, 1.8 μm); moreover, the mobile phase were consisted of 0.2% formic acid (A) and acetonitrile (B). In accordance with the findings, characterization of 120 chemical compounds was performed by liquid chromatography with mass spectrometry. The key constituents among them included ginsenosides (in Radix Ginseng), 16 triterpene carboxylic acids (in Poria), sesquiterpenes (in Rhizoma Atractylodis Macrocephalae), triterpenesaponins (in Glycyrrhizae Radix et Rhizoma Praeparata Cum Melle) as well as flavonoids (in Glycyrrhizae Radix et Rhizoma Praeparata Cum Melle) in Sijunzi decoction. This research developed the bases for prospective research associated with Sijunzi decoction, together with being expected to be useful to rapidly extract and characterize the constituents in other Traditional Chinese herbal formulations.
1. Introduction An extensive use of Traditional Chinese herbal formulation (TCMF) has been made across China, in addition to its neighbouring nations in the lab practices owing to its broad effectiveness, along with some adverse impacts [1]. The chemical constitutes of TCMFs usually possess complexity for the interaction impacts of the constituting herbs found in them. Moreover, the synergistic action of intricate chemical ingredients constitutes the substantial basis for their pharmacological impacts. Accordingly, developing a fast and efficient analysis approach to separate and identify intricate constitutions in TCMFs is quite necessary for the substantial basis of pharmacological research. Sijunzi decoction (SJZD) is termed as a renowned Chinese prescription, which was primarily recordation in the Song Dynasty. Four conventional herbal medicines, including Ginseng Radix et Rhizoma, Poria, Atractylodis Macrocephalae Rhizoma and Glycyrrhizae Radix et Rhizoma Praeparata Cum Melle make its key ingredients. In China, the use of SJZD has a long history to treat gastrointestinal issues, in addition to being capable of fighting nausea, vomiting, and diarrhoea in an effective manner. Currently, it has been revealed by the lab researches that SJZD, in combination with chemotherapy medicines, is quite ⁎
effective to treat and oesophagus cancer [1–3]. Nowadays, the bases of constituents' research of SJZD have been developed by some relative research works. Yang Liu et al. had performed the identification and characterization of ginsenoside, flavonoid and triterpenoid in SJZD with the help of high-performance liquid chromatography coupled with tandem mass spectrometry (LC/MSn) [2]. Nevertheless, the detection range, together with sensitivity of iontrap mass spectrometer, limited both the scope and precision of the findings. UHPLC coupled with Fourier transform ion cyclotron resonance mass spectrometry (FT-ICR-MS) brings forth an enormous benefit for quantitatively identifying intricate composition for its elevated responsiveness and precision. By usage of this technology, Yinan Wang et al. characterized 33 chemical compounds in Cortex Fraxini [4] and Ting Liu et al. characterized 174 chemical compounds in Gegenqinlian decoction as well as 107 prototypes and 67 metabolites in rats [5]. The reports demonstrated the efficient and practical of this method in chemical compounds and metabolites detection. Furthermore, it is capable of providing information about the fragmentation patterns of chemical compounds which could extend help to speculate the structure of other similar chemical constituents. In the current research work, a fast as well as efficient methodology
Corresponding authors. E-mail addresses: [email protected] (M. Zhao), [email protected] (C. Zhao).
https://doi.org/10.1016/j.jchromb.2018.04.009 Received 17 December 2017; Received in revised form 30 March 2018; Accepted 6 April 2018 Available online 07 April 2018 1570-0232/ © 2018 Elsevier B.V. All rights reserved.
Journal of Chromatography B 1086 (2018) 11–22
Z. Guan et al.
was developed to systematically characterize the chemical ingredients in SJZD with the help of UHPLC-FT-ICR-MS. The research is capable of laying the substantial bases, together with providing considerable information for the pharmacological research of SJZD.
of 0.4 g of dried powder was made using 5 mL menthol, followed by extraction with the help of ultrasonic with a period of 20 min.
2. Material and methods
The purchase of key ingredients including, Ginseng Radix et Rhizoma (batch number: 20,151,012; source: Jilin China), Poria (batch number: 13,120,201; source: Hunan China), Atractylodis Macrocephalae Rhizoma (batch number: 13,102,101; source: Zhejiang China) and Glycyrrhizae Radix et Rhizoma Praeparata Cum Melle (batch number: 13,112,501; source: Gansu China) was made from Guoda pharmacy (Shenyang, China), which followed identification by Professor Jingming Jia (Department of TCM, Shenyang Pharmaceutical University, Shenyang, China). The key source of the reference compounds (purity > 98%), including atractyloside A, schaftoside, ononin and glycyrrhizic acid was Shanghai yuanye Bio-Technology Co., Ltd. (Shanghai, China); ginsenoside Re, ginsenoside Rb1, formononetin, atractylenolide III and atractylenolide II were attained from the National Institute for the Control of Pharmaceutical and Biological Products (Beijing, China). In addition to that, Acetonitrile of HPLC grade, together with formic acid of LC-MS grade was attained from Fisher Scientific (Fair Lawn, NJ, USA), followed by attaining the purified water from Wahaha (Hangzhou, China).
The use of an Agilent 1260 UHPLC system (USA) was made in order to perform the chromatographic analysis. The use of a Universal XB C8 column (150 mm × 2.1 mm, 1.8 μm; Kromat, USA) having the column temperature of 45 °C was made. The mobile phase comprised 0.2% formic acid (A) and acetonitrile (B), which was conducted with gradient condition as hereunder: 2–2% (B) in 0 to 3 min, 2–22% (B) in 3 to 13 min, 22–70% (B) in 13 to 45 min. The setting of the flow rate at 0.20 mL·min−1 was ensured, together the injection volume amounting to be 2 μL. The use of a Bruker Solarix7.0 T FT-ICR-MS system (Bruker, Germany) as well as a Bruker Compass-Hystar workstation (Bruker, Germany) was made to carry out the mass spectra analysis. Ionization was carried out using positive as well as negative electrospray ionization (ESI) modes, followed by setting the optimized conditions as hereunder: nebulizer gas pressure, 4.0 bar; dry gas flow rate, 8 L·min−1; dry gas temperature, 200 °C; ion accumulation time, 0.15 s; time of flight, 0.6 ms; capillary voltage, 4.5 kV; and end plate offset, 500 V. Recording of the Full-scan mass spectrum data was performed between m/z 100 and 3000 amu. In respect of auto MS/MS mode, selection of both MS/MS Boost and MS/MS Isolation was made; moreover, range of the collision power lied between 10 eV and 30 eV as regards the MS/MS experimentations.
2.2. Preparation of SJZD for analysis
3. Results and discussion
In accordance with the documentary records of SJZD, four key constituting herbs that included Radix Ginseng (100 g), Poria (100 g), Rhizoma Atractylodis Macrocephalae (100 g), and Glycyrrhizae Radix et Rhizoma Praeparata Cum Melle (50 g) were crushed and made to form small pieces, followed by mixing and decocting two times in 3500 mL water for a period of 1 h each time in a glass flask [2,6]. Subsequently, mixing of the ensuing solution was done that followed drying with the help of lyophilization. Prior to the analysis, dissolution
The Fig. 1 reveals the base peak ion chromatograms (BPC) of SJZD in positive as well as negative ion modes, together with the respective compounds. The extract ion chromatograms (EIC) of each molecule weight were correspondingly attained for detecting the associated compound followed by presenting in Supporting information Fig. S1. Among the compounds identified, accurate identification of nine was carried out with the help of comparison with the retention time (tR) as well as the MS/MS data associated with the reference compounds in
2.3. Instrument and analytical conditions
2.1. Chemicals and materials
Fig. 1. The base peak ion chromatograms (BPC) of SJZD in both positive (A) and negative (B) ion modes and the corresponding compounds (C). 1-Atractyloside A, 2schaftoside, 3-ononin, 4-ginsenoside Re, 5-ginsenoside Rb1, 6-formononetin, 7-glycyrrhizic acid, 8-atractylenolide II, 9-atractylenolide III.
12
Journal of Chromatography B 1086 (2018) 11–22
Z. Guan et al.
Fig. 2. The chemical structures of chemical compounds detected in Radix Ginseng.
positive ion mode (Fig. 1). Moreover, the others were inferred by the retention times, in addition to molecular weight as well as MS/MS fragments. The use of the Bruker workstation was made for computing the molecular formulas of compounds through comparison with the identified molecular weights with the measured molecular weights, followed by accepting the error values being < 5.0 ppm. Further speculations of the layouts of the compounds were made using the MS/MS data. In aggregate, identification and characterization of 120 compounds were performed, whereas their layouts were presented in the Figs. 2-6. The presentation of MS/MS spectra of the usual compounds was provided in the Fig. 7, together with displaying their possible fragmentation pathways in the Fig. 8. Inferences of each and every ingredient were carried out with the help of the molecule formula and fragmentation pathways, followed by additional confirmation by reference to pertinent literature [7–16]. The Table 1 sheds light on the retention time, formula, molecular weight, calculated m/z, detected m/ z, error value and MS/MS data of the ingredients. Concrete illustration of the characterization of the ingredients in the individual lab materials was done as hereunder. 3.1. Characterization of constituents in Radix ginseng Saponins made the key active ingredients in Radix Ginseng, whereby, the majority of them were the glycosides of triterpenoid aglycones. In this study, tentative characterization of 34 triterpenoid saponins of Radix Ginseng in SJZD was carried out, followed by identifying 2 of them. Both Peak 4 and 5 in the Fig. 1C were ginsenoside Re as well as ginsenoside Rb1, correspondingly. Ginsenoside Rs5 is termed as a common ginsenoside, being less often stated in the literature, was put to use as an illustration for demonstrating the fragmentation pathways (Figs. 7A, 8A). In respect of the positive mode, the ion at m/z 831.48661 was deducted for being the adduct ion ([M + Na]+) and calculation of the formula was made as C44H72O13. The key fragment ions found in the MS/MS spectrum were at the m/z735.64263, 407.35689, 279.11181 as well as 181.07431. The ion at m/z 735.64263 revealed the loss of C2H2O3 (74 Da) from the precursor ion, whereas, the ion at m/z 407.35689 denoted the loss of C12H28O10 (328 Da) from m/z 735.64263. The ions at m/z 279.11181 as well as 181.07431 represented C11H18O8 and glucose, correspondingly. The characterization of other ingredient in Radix Ginseng was performed based on the fragmentation patterns and related literatures [7–10]. 3.2. Characterization of constituents in Poria 16 triterpene carboxylic acids made the key active ingredients in Poria [17]. In this research work, tentative characterization of 15 sixteen triterpene carboxylic acids of Poria in SJZD was performed. The use of Trametenolic acid, being a common composition of Poria was made as an instance for demonstrating the fragmentation pathways (Figs. 7B, 8B). In respect of the positive mode, inference of the ion at m/ z 421.33572 was made for being the key fragment ion, followed by indicating the loss of H2O2 (36 Da) from the precursor ion. The ion at m/z 2851.17113 in the MS/MS spectrum was an indication of the loss of C14H24 (192 Da) from the precursor ion. Characterization of the other constituents in Poria was carried out based on the fragmentation patterns and related literatures [11,12]. 3.3. Characterization of constituents in Rhizoma Atractylodis Macrocephalae Sesquiterpenes made the key active constitutes in Rhizoma Atractylodis Macrocephalae [18]. In this research work, tentative 13
Journal of Chromatography B 1086 (2018) 11–22
Z. Guan et al.
Fig. 3. The chemical structures of chemical compounds detected in Poria.
on the fragmentation patterns and related literatures [13,14].
characterization of 11 sesquiterpenes of Rhizoma Atractylodis Macrocephalae in SJZD was carried out, followed by precisely identifying three of them. Peak 1, 8 and 9 in the Fig. 1C were atractyloside A, atractylenolide II and atractylenolide, correspondingly. The use of Atractylenolide I, being a common composition of Rhizoma Atractylodis Macrocephalae was made as an instance for demonstrating the fragmentation pathways (Figs. 7C, 8C). In respect of the positive mode, inference of the ion at m/z 233.15386 was performed for being the adduct ion ([M + H]+), followed by calculation of the formula as C15H20O2. The key fragment ions found in the MS/MS spectrum were at the m/z 215.14072, 187.14559, 177.08796 as well as 145.10055. The ion at m/z 215.14072 was an indication of the loss of H2O (18 Da) from the precursor ion. The ions at m/z 187.14559 as well as 177.08796 were the indications of the loss of CO (28 Da) and C3H2 (38 Da) from m/ z 215.14072. Moreover, the ions at m/z 145.10055 suggested the loss of CH4O (32 Da) from m/z 177.08796. Characterization of the other ingredients in Rhizoma Atractylodis Macrocephalae was performed based
3.4. Characterization of constituents in Glycyrrhizae Radix et Rhizoma Praeparata Cum Melle Triterpenesaponins, together with several kinds of flavonoids, made the key active constituents in Glycyrrhizae Radix et Rhizoma Praeparata Cum Melle [15]. In this research work, tentative characterization of 60 ingredients of Glycyrrhizae Radix et Rhizoma Praeparata Cum Melle in SJZD was performed, followed by precise identification of 4 among them. Peak 2, 3, 6 and 7 presented in the in Fig. 1C represented schaftoside, ononin, formononetin and glycyrrhizic acid, correspondingly. The use of Liquiritigenin, being a common flavonoids composition of Glycyrrhizae Radix et Rhizoma Praeparata Cum Melle was made as an illustration for demonstrating the fragmentation pathways (Fig. 7D, 8D). In respect of the positive mode, inference of the ion at m/z 257.08089 was performed for being the adduct ion
14
Journal of Chromatography B 1086 (2018) 11–22
Z. Guan et al.
Fig. 4. The chemical structures of chemical compounds detected in Rhizoma Atractylodis Macrocephalae.
chemical ingredients existing in SJZD. The findings revealed the fact that ginsenosides (in Radix Ginseng), sixteen triterpene carboxylic acids (in Poria), Sesquiterpenes (in Rhizoma Atractylodis Macrocephalae), triterpenesaponins (in Glycyrrhizae Radix et Rhizoma Praeparata Cum Melle) as well as flavonoids (in Glycyrrhizae Radix et Rhizoma Praeparata Cum Melle) made the key ingredients existing in SJZD, followed by the provision of the particular data. This research provided more information of chemical compounds in SJZD than previous research in this field. Therefore, good amount of substantial foundation, together with considerable amount of information has been provided by this research work in respect of the pharmacological research of SJZD. This research together with other similar papers could be scientific proof for the chemical ingredients characterization on this method of other TCMFs. However, in this research, the chemical structures of compounds were speculated based on the precise molecule formula of MS and MS/ MS information and fragmentation pathways, followed by additional confirmation by reference to pertinent literature. The identification of structure was limited to known compounds only, which is also the limitation of chemical structure identification by mass spectrometry. Thus, the advantages of mass spectrometry identification were rapid and effective, for the more precise identification, assistance in other technologies such as nuclear magnetic resonance (NMR) and single crystal X-ray diffraction (SXRD) were required. Supplementary data to this article can be found online at https:// doi.org/10.1016/j.jchromb.2018.04.009.
([M + H]+), followed by the calculation of the formula as C15H13O4. The key fragment ions found in the MS/MS spectrum were at the m/z 147.04216, 137.02175 as well as 119.04821. The ion at m/z 147.04216 as well as137.02175 suggested the loss of C6H6O2 (110 Da) and C8H8O (120 Da) from the precursor ion, correspondingly. The ion at m/z 119.04821 denotes the loss of CO (28 Da) from m/z 147.04216. Uralsaponin N was used as a common triterpenesaponins composition of Glycyrrhizae Radix et Rhizoma Praeparata Cum Melle as an illustration for demonstrating the fragmentation pathways (Figs. 7E, 8E). In the positive mode, the ion at m/z 839.40598 was inferenced to be the adduct ion ([M + H]+), followed by the calculation of the formula as C42H63O17. The key fragment ions found in the MS/MS spectrum were at the m/z 663.37644, 487.32988 as well as 179.05516. The ion at m/z 663.37644 suggested the loss of C6H9O6 (178 Da) from the precursor ion which indicated the breaking of glycosidic bond. The ion at m/z 179.05516 was calculated as C6H10O6 as another fragment ion produced by the breaking of glycosidic bond. The ion at m/z 487.32988 suggested the loss of C6H9O6 (178 Da) from the precursor ion m/z 663.37644. Characterization of the other ingredients in Glycyrrhizae Radix et Rhizoma was performed based on the fragmentation patterns and related literatures [15,16]. 4. Conclusions A swift and straightforward methodology was performed to systematically characterize the chemical constituents found in SJZD with the help of UHPLC-FT-ICR-MS. This methodology brought forth shorttime as well as effective separation, together with detection of the
15
Journal of Chromatography B 1086 (2018) 11–22
Z. Guan et al.
Fig. 5. The chemical structures of flavonoids detected in Glycyrrhizae Radix et Rhizoma Praeparata Cum Melle.
16
Journal of Chromatography B 1086 (2018) 11–22
Z. Guan et al.
Fig. 6. The chemical structures of triterpenesaponins detected in Glycyrrhizae Radix et Rhizoma Praeparata Cum Melle.
17
Journal of Chromatography B 1086 (2018) 11–22
Z. Guan et al.
Fig. 7. The MS/MS spectra of the typical compounds. A-Ginsenoside Rs5, B-trametenolic acid, C-atractylenolide I, D-liquiritigenin, E-uralsaponin N.
18
Journal of Chromatography B 1086 (2018) 11–22
Z. Guan et al.
Fig. 8. The possible fragmentation pathways of the typical compounds. A-Ginsenoside Rs5, B-trametenolic acid, C-atractylenolide I, D-liquiritigenin, E-uralsaponin N.
19
Rt (min)
35.77 36.92 29.67 28.55 40.61 29.63 24.65 33.41 19.88 19.84 28.42 25.39 21.60 32.11 29.27 33.76 31.81 30.94 24.73 23.57 26.71 22.56 25.39 28.25 31.09 27.16 21.62 41.50 28.05 24.43 28.29 32.36 30.79 28.57 31.34
26.27 26.27 28.17 26.97 30.40 30.65
30.16
34.11
20.21 37.51 28.53 19.98 13.97 21.32
No.
1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35
36 37 38 39 40 41
42
43
44 45 46 47 48 49
Dehydroprotopanaxadiol II Dehydroprotopanaxatriol I Dehydroprotopanaxatriol II Panaxadione Ginsenoside Rk2 Ginsenoside Rk3 Ginsenoside Rh4 Compound K Ginsenoside F1 Ginsenoside Rh1 Ginsenoside ST2 Ocotillol derivative 3a Ginsenoside Km Ginsenoside Rs6 Compound Y Ginsenoside Rg6 Ginsenoside Rk1 Ginsenoside Rz1 Ginsenoside Rg5 Ginsenoside Rg3 Ginsenoside SL2 12,23-Epoxyginsenoside Rg1 Ginsenoside Rf Ginsenoside ORh1 Chikusetsusaponin IVa Floralginsenoside F Floralginsenoside E Ginsenoside Rs5 C-Y2 Ginsenoside Rg1 Ginsenoside Re Ginsenoside Ro Floralginsenoside Kb Ginsenoside Rb1 3β,16α-Dihydroxy-lanostane −7.9(11),24-Trien-21-oic acid Dehydrotrametenolic acid 16α-Hydroxytrametenolic acid Trametenolic acid Poricoic acid B 16-Deoxyporicoic acid B 3β-Hydroxy-16α-acetyloxy-lanosta −7.9(11),24-trien-21-oic acid 3-O-acetyl-16α-hydroxy -dehydrotrametenolic acid 3-O-acetyl-16α-hydroxytrametenolic acid Me dehydroeburicoate 6α-Hydroxydehydropachymic acid Poricoic acid DM Poricoic acid A Ganoderic acid B 3β,16α-Diacetoxy-24-(hydroxymethyl) lanost-8-en-21-oic acid
Identification
Monoisotopic mass (m/z) 443.38836 459.38327 459.38327 511.33940 605.44118 621.43797 621.43797 623.45175 639.44666 639.44666 655.44157 655.44157 655.44157 663.44666 755.49400 767.49400 767.49400 767.49400 767.49400 785.50457 799.48383 799.48383 801.49948 803.50701 817.43448 817.49440 817.49440 831.48651 845.49041 845.49041 947.55739 957.50536 965.49628 1153.60114 453.33632 455.35197 455.35197 457.36762 467.31559 469.33124 495.34689 495.34689 515.37310 521.33915 541.35346 529.35237 537.29768 555.27186 589.40988
Formula C30H51O2 C30H51O3 C30H51O3 C30H48O5Na C36H61O7 C36H61O8 C36H61O8 C36H63O8 C36H63O9 C36H63O9 C36H63O10 C36H63O10 C36H63O10 C38H63O9 C41H71O12 C42H71O12 C42H71O12 C42H71O12 C42H71O12 C42H73O13 C42H71O14 C42H71O14 C42H73O14 C44H76O10K C42H66O14Na C42H73O15 C42H73O15 C44H72O13Na C43H73O16 C43H73O16 C43H83O18 C48H77O19 C46H77O21 C55H93O25 C30H45O3 C30H47O3 C30H47O3 C30H49O3 C30H43O4 C30H45O4 C32H47O4 C32H47O4 C32H51O5 C32H50O3K C33H49O6 C32H49O6 C31H46O5K C30H44O7K C35H57O7
Table 1 The results of chemical constituent characterization of SJZD by FT-ICR-MS.
20 521.33716 541.35556 529.35323 537.29581 555.27363 589.41266
515.37173
495.34708
455.35238 455.35238 457.36812 467.31580 469.33261 495.34754
443.38986 459.38375 459.38483 511.34040 605.44157 621.43768 621.43797 623.45176 639.44633 639.44632 655.44311 655.44151 655.44142 663.44666 755.49483 767.49463 767.49621 767.49531 767.49425 785.50339 799.48337 799.48303 801.49803 803.50802 817.43492 817.49388 817.49342 831.48661 845.48747 845.48718 947.56099 957.50368 965.49370 1153.59692 453.33710
Detected mass (m/z)
−0.40
[M − H2O + H]+
[M + K]+ [M − H]− [M + H]+ [M + K]+ [M + K]+ [M + H]+
[M + H]
3.82 −3.88 −1.64 3.48 −3.19 −4.72
2.66
395.36772, 437.33189, 421.33572, 409.28485, 397.31662, 437.33282,
−0.89 −0.89 −1.10 −0.46 −2.93 −1.32 [M + H]+ [M − H2O + H]+ [M + H]+ [M − H2O + H]+ [M + H]+ [M − H2O + H]+
+
423.36375, 409.38346, 423.36409, 443.38708, 443.38874, 459.38368, 409.38312, 461.39789, 603.41985, 603.42033, 493.38126, 493.38315, 506.39120, 409.38422, 461.39637, 603.42658, 443.38763, 443.38900, 587.43633, 749.48820, 491.36956, 475.37688, 621.43579, 477.38776, 631.38351, 493.39046, 493.39607, 735.64263, 475.38546, 475.38796, 621.42643, 631.38258, 447.35577, 783.48952, 435.31284,
−3.40 −1.05 −3.40 −1.96 −0.65 −2.54 −3.02 −0.03 0.52 0.53 −2.35 0.09 0.23 0.00 −1.09 −0.82 −2.87 −1.70 −0.31 1.50 0.57 1.01 1.81 −1.26 −0.55 0.63 1.19 −0.11 3.47 3.82 −3.80 1.75 2.68 3.66 −1.73 [M + H]+ [M + H]+ [M + H]+ [M + Na]+ [M + H]+ [M + H]+ [M + H]+ [M + H]+ [M + H]+ [M + H]+ [M + H]+ [M + H]+ [M + H]+ [M + H]+ [M + H]+ [M + H]+ [M + H]+ [M + H]+ [M + H]+ [M + H]+ [M + H]+ [M + H]+ [M + H]+ [M + K]+ [M + Na]+ [M + H]+ [M + H]+ [M + Na]+ [M + HCOOH-H]− [M + HCOOH-H]− [M + H]+ [M + H]+ [M + HCOOH-H]− [M + HCOOH-H]− [M − H2O + H]+ 299.26112 297.25031 215.17499 277.20385 283.23173 297.25884
109.10256 69.07763 109.10193 69.07015 109.10267 109.10304 69.07651 165.06845 163.05846 163.05694 163.05773 163.06004 163.06129 69.07641 296.09754 147.06394 109.10288 163.05773 407.36994 441.3765 310.11906 165.06757 163.05988 290.16350 163.06396 163.05481 326.11348 287.06043 308.11691 179.05378 179.05255 177.03002 179.05482 341.10843 309.25784
467.38808, 501.35669, 441.29522, 426.31107, 481.28831, 505.38933,
409.37226, 447.31274, 397.30047, 381.30452, 325.20193, 425.37127,
P P P P P P
P
P
P P P P P P
G G G G G G G G G G G G G G G G G G G G G G G G G G G G G G G G G G P
Source
(continued on next page)
299.26462 403.32992 279.20338 279.20475 297.25088 297.25650
473.35293, 437.33901, 393.34220
435.31290, 391.32978, 295.23088
325.28528, 393.34681, 251.17113, 365.29466, 353.32037, 393.34113,
407.36852, 121.10114, 407.36769, 359.29484, 407.36706, 407.36829, 121.10288, 429.40112, 457.36497, 565.37326, 428.40582, 459.38149, 443.41207, 121.09831, 429.40128, 459.38674, 407.36596, 411.39626, 425.37169, 605.44168, 443.71847, 445.39620, 457.36484, 429.40254, 455.35669, 444.73498, 444.72972, 407.35689, 427.40177, 411.40398, 441.36383, 455.35381, 341.03776, 603.42612, 391.32005,
MS/MS (m/z)
ppm
Ion mode
Z. Guan et al.
Journal of Chromatography B 1086 (2018) 11–22
Rt (min)
34.66 21.04 37.90 34.66 23.87 32.63 42.52 24.83 16.74 15.50 14.91 25.26 20.81 24.43 31.98 19.19 22.84 24.52 32.26 36.13 35.38 33.28 35.44 34.26 2.65 33.86 31.98 29.58 39.17 37.83 35.71 27.64 18.16
27.39 35.71 18.08 20.58 17.14 31.38 24.43 20.30 22.75 17.38 20.30 41.52 20.22 20.60 23.64 18.68 18.98 16.14 19.88 17.64
No.
50 51 52 53 54 55 56 57 58 59 60 61 62 63 64 65 66 67 68 69 70 71 72 73 74 75 76 77 78 79 80 81 82
83 84 85 86 87 88 89 90 91 92 93 94 95 96 97 98 99 100 101 102
Table 1 (continued)
Atractylenolide II Atractylentrid Atractylenolide I Atractylenolide III Chrysin Selina-4(14),7(11)-dien-8-one Dibutyl phthalate Luteolin Icariside D1 Atractyloside A Atractyloside B Pinocembrin Liquiritigenin Formononetin Echinatin Vestitol Naringenin Licochalcone B Glabrene Glabrone Licochalcone A Licoflavonol 3′-O-methyl glabridin 4′-O-methyl glabridin Kumatakenin Glicoricone Licoarylcoumarin Glycycoumarin Glyasperins D Glycyrin Licoricone Licopyranocoumarin 7-Hydroxy-2-(4-hydroxyphenyl)-6,8-bis (3-methyl-2-buten-1-yl)-2,3-dihydro-4Hchromen-4-one Kanzonol M Kanzonol O 6,8-Diprenylgenistein Isoliquiritin Liquiritin Kanzonol F Ononin Vitexin Choerospondin Licoagroside B Trifolirhizin Glycyrrhetinic acid Licoagroside A liquiritin Apioside Isoliquiritin apioside Schaftoside Isoschaftoside Violanthin Isoviolanthin Vicenin-2
Identification
21 399.18022 405.13086 407.18530 419.13366 419.13366 421.20095 431.13366 433.11292 433.11402 433.13405 447.12857 471.34689 491.11950 551.17592 551.17592 565.15518 565.15518 579.17083 579.17083 595.16575
231.13796 233.11722 233.15361 249.14852 253.05063 257.13022 279.15909 285.04046 439.15747 471.22007 495.24470 257.08084 257.08084 269.08084 271.09649 271.11214 273.07575 285.07685 321.11323 335.09250 337.14453 355.11761 355.15400 355.15400 359.07724 369.13326 369.13326 369.13326 371.18530 383.14891 383.14891 385.12818 393.20604
C15H19O2 C14H17O3 C15H21O2 C15H21O3 C15H9O4 C15H22OK C16H23O4 C15H9O6 C19H28O10Na C21H36O10Na C22H39O12 C15H13O4 C15H13O4 C16H13O4 C16H15O4 C16H17O4 C15H13O5 C16H13O5 C20H17O4 C20H15O5 C21H21O4 C20H19O6 C21H23O5 C21H23O5 C18H15O8 C21H21O6 C21H21O6 C21H21O6 C22H26O5 C22H23O6 C22H23O6 C21H21O7 C25H29O4
C23H27O6 C22H22O6Na C25H27O5 C21H23O9 C21H23O9 C26H29O5 C22H23O9 C21H21O10 C21H21O10 C18H25O12 C22H23O10 C30H47O4 C23H23O12 C26H31O13 C26H31O13 C26H29O14 C26H29O14 C27H31O14 C27H31O14 C27H31O15
Monoisotopic mass (m/z)
Formula
399.17838 405.13141 407.18672 419.13391 419.13375 421.20184 431.13320 433.11297 433.11249 433.13396 447.12865 471.34787 491.11738 551.17559 551.17577 565.15456 565.15500 579.17010 579.17031 595.16480
231.13815 233.11680 233.15386 249.14875 253.05018 257.13052 279.15956 285.03959 439.15746 471.21966 495.24294 257.08103 257.08089 269.08117 271.09637 273.11218 273.07594 285.07627 321.11278 335.09185 337.14387 355.11809 355.15440 355.15431 359.07639 369.13363 369.13357 369.13426 371.18615 383.14945 383.14927 385.12846 393.20743
Detected mass (m/z)
4.61 −1.35 −3.49 −0.60 −0.22 −2.11 1.06 −0.11 3.52 0.21 −0.16 −2.09 4.32 0.59 0.27 1.10 0.33 1.27 0.90 1.60
[M + H]+ [M + Na]+ [M + H]+ [M + H]+ [M + H]+ [M + H]+ [M + H]+ [M + H]+ [M − H]− [M + H]+ [M + H]+ [M + H]+ [M − H]− [M + H]+ [M + H]+ [M + H]+ [M + H]+ [M + H]+ [M + H]+ [M + H]+
367.15793, 375.13794, 337.11593, 309.10743, 282.11531, 389.17549, 399.11572, 269.04531, 282.11643, 309.10225, 285.07128, 339.26538, 327.07693, 417.12641, 417.12548, 446.11564, 446.11986, 460.12586, 460.12536, 476.12561,
213.12351, 185.13547, 215.14072, 231.13685, 209.06399, 202.72643, 205.08378, 201.01564, 285.13978, 287.18664, 287.19446, 152.01593, 147.04216, 237.06352, 239.07549, 241.09571, 152.01597, 255.07891, 270.09511, 281.06382, 307.13591, 299.06137, 323.13596, 307.13521, 345.06571, 313.07536, 353.10593, 337.11578, 339.16597, 351.12591, 327.09253, 353.10521, 323.13571,
−0.86 1.80 −1.10 −0.92 1.79 −1.16 −1.69 3.07 0.01 0.87 3.55 −0.75 −0.23 −1.26 0.44 −0.15 −0.70 2.04 1.40 1.92 1.97 −1.34 −1.13 −0.88 2.37 −0.98 −0.84 −2.69 −2.30 −1.40 −0.93 −0.73 −3.53 [M + H] [M + H]+ [M + H]+ [M + H]+ [M − H]− [M + K]+ [M + H]+ [M − H]− [M + Na]+ [M + Na]+ [M + HCOOH-H]− [M + H]+ [M + H]+ [M + H]+ [M + H]+ [M + H]+ [M + H]+ [M − H]− [M − H]− [M − H]− [M − H]− [M + H]+ [M + H]+ [M + H]+ [M + HCOOH-H]− [M + H]+ [M + H]+ [M + H]+ [M + H]+ [M + H]+ [M + H]+ [M + H]+ [M + H]+
175.06651 83.04337 145.10055 187.14653 107.01556 105.06856 57.06455 107.01345 79.05661 139.14358 137.14063 104.06359 119.04821 136.02586 121.03782 123.04827 120.06783 165.06538 214.10529 136.02593 217.12435 165.02596 285.11495 269.12734 328.06134 136.02543 301.07541 206.09162 303.12483 315.09157 312.06586 162.03716 138.03632
Gly Gly Gly Gly Gly Gly Gly Gly Gly Gly Gly Gly Gly Gly Gly Gly Gly Gly Gly Gly
A A A A A A A A A A A Gly Gly Gly Gly Gly Gly Gly Gly Gly Gly Gly Gly Gly Gly Gly Gly Gly Gly Gly Gly Gly Gly
Source
(continued on next page)
338.15431, 331.12769 353.11682, 337.12583 268.04196, 155.03697 281.10593, 255.07495 255.07461, 136.02561 367.15319, 353.14128 267.07593, 122.04536 163.06321,154.03257 271.06593, 152.01479 147.05422, 127.02467 229.08474, 149.02177 189.16722, 137.13835 164.03532, 148.03643 255.07631, 136.02758 255.07953, 137.02497 431.10298, 401.09589 431.10358, 401.09658 431.10581, 415.10692 431.10857, 415.10486 431.10592, 270.05391
185.12567, 153.08055, 187.14559, 215.13995, 143.05648, 178.96543, 149.02367, 133.02614, 123.07536, 183.13114, 183.14758, 124.02671, 137.02175, 148.05293, 149.06349, 149.06493, 124.02641, 193.05761, 253.09137, 267.07936, 245.12961, 271.05284, 301.11679, 285.11357, 344.06257, 299.06351, 337.11593, 301.07152, 309.15674, 321.11539, 315.08961, 222.09143, 254.06395,
MS/MS (m/z)
+
ppm
Ion mode
Z. Guan et al.
Journal of Chromatography B 1086 (2018) 11–22
Journal of Chromatography B 1086 (2018) 11–22
3.18 −0.05 −0.58 0.85 0.93 −2.13 1.10 0.65 −0.11 [M − H]− [M + H]+ [M + Na]+ [M + H]+ [M + H]+ [M + Na]+ [M + Na]+ [M + H]+ [M + H]+ 849.35234 881.41659 887.40409 897.41069 955.48882 979.48939 979.48622 985.46325 1027.47457 849.35504 881.41654 887.40357 897.41146 955.48971 979.48730 979.48730 985.46389 1027.47445 27.85 28.47 30.67 27.37 26.97 29.83 32.38 27.84 27.92 112 113 114 115 116 117 118 119 120
Uralsaponin E 22β-Acetoxyl-glycyrrhizin 22β-Acetoxyl-glycyrrhaldehyde Uralsaponin F Uralsaponin T Yunganoside A1 Yunganoside B1 Licorice-saponin A3 Uralsaponin X
C42H57O18 C44H65O18 C44H64O17Na C44H65O19 C48H75O19 C48H76O19Na C48H76O19Na C48H73O21 C50H75O22
694.21268 697.21272 807.41569 821.39689 823.41322 825.42596 831.41360 839.40690 694.21303 697.21270 807.41615 821.39541 823.41106 825.42671 831.41374 839.40598 C35H36O14N C35H37O15 C42H63O15 C42H61O16 C42H63O16 C42H65O16 C42H64O15Na C42H63O17 Licorice glycoside E Licorice glycoside B Uralsaponin W Uralsaponin O Glycyrrhizic acid Uralsaponin P Licorice-saponin B2 Uralsaponin N 26.03 24.48 34.83 30.35 31.38 26.29 32.53 30.47 103 104 105 106 107 108 109 111
“The authors declare that there are no conflicts of interest”. Acknowledgements The authors gratefully acknowledge the facilities and assistance provided by Dr. Fei Han of the Bruker Corporation. G represented Ginseng Radix et Rhizoma; P represented Poria; A represented Atractylodis Macrocephalae Rhizoma; Gly represented Glycyrrhizae Radix et Rhizoma Praeparata Cum Melle.
Gly Gly Gly Gly Gly Gly Gly Gly Gly 179.04641 179.04753 179.04966 177.06586 179.04616 149.07233 149.07347 179.04862 149.08465 485.31611, 529.34658, 513.34358, 545.33421, 458.35522, 459.37546, 459.37235, 472.33458, 529.34414,
Gly Gly Gly Gly Gly Gly Gly Gly 294.08615, 257.07166 297.08341, 257.07246 475.37620, 181.06385 491.36486, 181.06344 471.32977, 179.04623 487.33574, 165.06255 238.17058, 224.19570 487. 32,988.
440.14134, 443.14235, 653.41902, 669.41376, 647.36821, 663.36548, 413.37584, 663.37644, 179.05516 661.34962, 705.37623, 689.37723, 721.36447, 779.44623, 811.43655, 811.43913, 809.42963, 705.37552, 0.51 −0.04 0.56 −1.80 −2.74 0.91 0.17 −1.10 [M + H] [M + H]+ [M + H]+ [M + H]+ [M + H]+ [M + H]+ [M + Na]+ [M + H]+
MS/MS (m/z)
+
Ion mode Detected mass (m/z) Monoisotopic mass (m/z) Formula Identification Rt (min) No.
Table 1 (continued)
Conflicts of interest
ppm
Source
Z. Guan et al.
Funding This work was supported by the National Natural Science Foundation of China (NSFC: 81503035). This work was supported by the Shenyang Pharmaceutical University Young Teachers' Career Development Program. References [1] Kang An, Guo Jin-rui, Zhang Zhen, Wang Xiao-long, Simultaneous quantification of ten active components in traditional Chinese formula Sijunzi decoction using a UPLC-PDA method, J. Anal. Methods Chem. 570359 (2014) 2–8. [2] Yang Liu, Junshan Yang, Zongwei Cai, Chemical investigation on Sijunzi decoction and its two major herbs Panax ginseng and Glycyrrhiza uralensis by LC/MS/MS, J. Pharm. Biomed. Sci. 41 (2006) 1642−1647. [3] Zhibo Guan, Juan Wu, Cancan Wang, Fang Zhang, Yinan Wang, Miao Wang, Min Zhao, Chunjie Zhao, Investigation of the preventive effect of Sijunzi decoction on mitomycin C induced immunotoxicity in rats by 1H NMR and MS-based untargeted metabolomic analysis, J. Ethnopharmacol. 210 (2018) 179–191. [4] Yinan Wang, Fei Han, Aihua Song, Miao Wang, Min Zhao, Chunjie Zhao, Rapid characterization of the chemical constituents of Cortex Fraxini by homogenate extraction followed by UHPLC coupled with Fourier transform ion cyclotron resonance mass spectrometry and GC–MS, J. Sep. Sci. 00 (2016) 1–10. [5] Ting Liu, Xiumin Tian, Zhaoqin Li, Fei Han, Bin Ji, Yunli Zhao, Zhiguo Yu, Metabolic profiling of Gegenqinlian decoction in rat plasma, urine, bile and feces after oral administration by ultra high performance liquid chromatography coupled with Fourier transform ion cyclotron resonance mass spectrometry, J. Chromatogr. B 1079 (2018) 69–84. [6] Qiuxiang Xu, Hongli Yu, Hao Wu, Fenqiang You, Yunhan Zhu, Lijuan Tang, Pharmacological comparison between traditional decoction and machine decoction of Sijunzi Tang, Chin. J. Exp. Tradit. Med. Formulae. 19 (2013) 250–255. [7] Byong-Kyu Shin, Sung Won Kwon, Jeong Hill Park, Chemical diversity of ginseng saponins from Panax ginseng, J. Ginseng Res. 39 (2015) 287−298. [8] Zai-Qun Liu, Chemical insights into ginseng as a resource for natural antioxidants, Chem. Rev. 112 (2012) 3329−3355. [9] JaeWon Lee, Ram Choi, Young-Chang Kim, Doo Jin Choi, Young-Seob Lee, GeumSoog Kim, Nam-In Baek, Seung-Yu Kim, Dae Young Lee, Comprehensive profiling and quantification of ginsenosides in the root, stem, leaf, and berry of Panax ginseng by UPLC-QTOF/MS, Molecules 22 (2017) 2147. [10] Lei Zhang, Qi-Le Zhou, Xiu-Wei Yang, Determination of the transformation of ginsenosides in Ginseng Radix et Rhizoma during decoction with water using ultrafast liquid chromatography coupled with tandem mass spectrometry, J. Sep. Sci. 41 (2018) 1039–1049. [11] Bing Xia, Yan Zhou, Hong Sheng Tan, Sheng Ding Li, Xu Hong Xi, Advanced ultraperformance liquid chromatography–photodiode array–quadrupole time-of-flight mass spectrometric methods for simultaneous screening and quantification of triterpenoids in Poria cocos, Food Chem. 152 (2014) 237–244. [12] José-Luis Ríos, Chemical constituents and pharmacological properties of Poria cocos, Planta Med. 77 (2011) 681–691. [13] Guo-Shun Shan, Liang-Xiao Zhang, Qi-Miao Zhao, Hong-Bin Xiao, Rong-Jie Zhuo, Gang Xua, Hong Jiang, Xian-Min You, Tian-Zhu Jia, Metabolomic study of raw and processed Atractylodes macrocephala Koidz by LC–MS, J. Pharm. Biomed. Anal. 98 (2014) 74–84. [14] Xu Yangyang, Hao Cai, Gang Cao, Yu Duan, Pei Ke, Tu Sicong, Jia Zhou, Xie Li, Dongdong Sun, Jiayu Zhao, Jing Liu, Xiaoqi Wang, Lin Shen, Profiling and analysis of multiple constituents in Baizhu Shaoyao San before and after processing by stirfrying using UHPLC/Q-TOF-MS/MS coupled with multivariate statistical analysis, J. Chromatogr. B 1083 (2018) 110–123. [15] Ci-Hai Zhang, Yue Yu, Yi-Zeng Liang, Xiao-Qing Chen, Purification, partial characterization and antioxidant activity ofpolysaccharides from Glycyrrhiza uralensis, Int. J. Biol. Macromol. 79 (2015) 681–686. [16] Wei Song, Longlong Si, Shuai Ji, Han Wang, Xiao-mei Fang, Yu Li-yan, Ren-yong Li, Li-na Liang, Demin Zhou, Min Ye, Uralsaponins M−Y, antiviral triterpenoid saponins from the roots of Glycyrrhiza uralensis, J. Nat. Prod. 77 (2014) 1632–1643. [17] Haruo Nukaya, Hirokazu Yamashiro, Hirotatsu Fukazawa, Hitoshi Ishida, Kuniro Tsuji, Isolation of inhibitors of TPA-induced mouse ear edema from Hoelen, Poria cocos, Chem. Pharm. Bull. 44 (1996) 847–849. [18] Qinhua Chen, Hongsheng He, Peng Li, Jun Zhu, Min Xiong, Identification and quantification of atractylenolide I and atractylenolide III in Rhizoma Atractylodes Macrocephala by liquid chromatography–ion trap mass spectrometry, Biomed. Chromatogr. 27 (2013) 699–707.
22
Contents lists available at ScienceDirect
Journal of Chromatography B journal homepage: www.elsevier.com/locate/jchromb
Review
Rapid characterization of the chemical constituents of Sijunzi decoction by UHPLC coupled with Fourier transform ion cyclotron resonance mass spectrometry ⁎
Zhibo Guana, Miao Wangb, Yi Caia, Hongmei Yanga, Min Zhaoa, , Chunjie Zhaoa, a b
T
⁎
School of Pharmacy, Shenyang Pharmaceutical University, Wenhua Road 103, Shenyang, Liaoning Province, China School of Life Science and Biopharmaceutics, Shenyang Pharmaceutical University, Wenhua Road 103, Shenyang, Liaoning Province, China
A R T I C LE I N FO
A B S T R A C T
Keywords: Sijunzi decoction Poria Atractylodis Macrocephalae Rhizoma Fourier transform ion cyclotron resonance Chemical constituents
Sijunzi decoction, a renowned Chinese prescription has long been utilized to treat gastrointestinal problems. In the context of this research work, the use of Ultra high performance liquid chromatography combined with Fourier transform ion cyclotron resonance mass spectrometry was made to separate and characterize the components of Sijunzi decoction. The performance of Liquid chromatography was carried out on a C8 column (150 mm × 2.1 mm, 1.8 μm); moreover, the mobile phase were consisted of 0.2% formic acid (A) and acetonitrile (B). In accordance with the findings, characterization of 120 chemical compounds was performed by liquid chromatography with mass spectrometry. The key constituents among them included ginsenosides (in Radix Ginseng), 16 triterpene carboxylic acids (in Poria), sesquiterpenes (in Rhizoma Atractylodis Macrocephalae), triterpenesaponins (in Glycyrrhizae Radix et Rhizoma Praeparata Cum Melle) as well as flavonoids (in Glycyrrhizae Radix et Rhizoma Praeparata Cum Melle) in Sijunzi decoction. This research developed the bases for prospective research associated with Sijunzi decoction, together with being expected to be useful to rapidly extract and characterize the constituents in other Traditional Chinese herbal formulations.
1. Introduction An extensive use of Traditional Chinese herbal formulation (TCMF) has been made across China, in addition to its neighbouring nations in the lab practices owing to its broad effectiveness, along with some adverse impacts [1]. The chemical constitutes of TCMFs usually possess complexity for the interaction impacts of the constituting herbs found in them. Moreover, the synergistic action of intricate chemical ingredients constitutes the substantial basis for their pharmacological impacts. Accordingly, developing a fast and efficient analysis approach to separate and identify intricate constitutions in TCMFs is quite necessary for the substantial basis of pharmacological research. Sijunzi decoction (SJZD) is termed as a renowned Chinese prescription, which was primarily recordation in the Song Dynasty. Four conventional herbal medicines, including Ginseng Radix et Rhizoma, Poria, Atractylodis Macrocephalae Rhizoma and Glycyrrhizae Radix et Rhizoma Praeparata Cum Melle make its key ingredients. In China, the use of SJZD has a long history to treat gastrointestinal issues, in addition to being capable of fighting nausea, vomiting, and diarrhoea in an effective manner. Currently, it has been revealed by the lab researches that SJZD, in combination with chemotherapy medicines, is quite ⁎
effective to treat and oesophagus cancer [1–3]. Nowadays, the bases of constituents' research of SJZD have been developed by some relative research works. Yang Liu et al. had performed the identification and characterization of ginsenoside, flavonoid and triterpenoid in SJZD with the help of high-performance liquid chromatography coupled with tandem mass spectrometry (LC/MSn) [2]. Nevertheless, the detection range, together with sensitivity of iontrap mass spectrometer, limited both the scope and precision of the findings. UHPLC coupled with Fourier transform ion cyclotron resonance mass spectrometry (FT-ICR-MS) brings forth an enormous benefit for quantitatively identifying intricate composition for its elevated responsiveness and precision. By usage of this technology, Yinan Wang et al. characterized 33 chemical compounds in Cortex Fraxini [4] and Ting Liu et al. characterized 174 chemical compounds in Gegenqinlian decoction as well as 107 prototypes and 67 metabolites in rats [5]. The reports demonstrated the efficient and practical of this method in chemical compounds and metabolites detection. Furthermore, it is capable of providing information about the fragmentation patterns of chemical compounds which could extend help to speculate the structure of other similar chemical constituents. In the current research work, a fast as well as efficient methodology
Corresponding authors. E-mail addresses: [email protected] (M. Zhao), [email protected] (C. Zhao).
https://doi.org/10.1016/j.jchromb.2018.04.009 Received 17 December 2017; Received in revised form 30 March 2018; Accepted 6 April 2018 Available online 07 April 2018 1570-0232/ © 2018 Elsevier B.V. All rights reserved.
Journal of Chromatography B 1086 (2018) 11–22
Z. Guan et al.
was developed to systematically characterize the chemical ingredients in SJZD with the help of UHPLC-FT-ICR-MS. The research is capable of laying the substantial bases, together with providing considerable information for the pharmacological research of SJZD.
of 0.4 g of dried powder was made using 5 mL menthol, followed by extraction with the help of ultrasonic with a period of 20 min.
2. Material and methods
The purchase of key ingredients including, Ginseng Radix et Rhizoma (batch number: 20,151,012; source: Jilin China), Poria (batch number: 13,120,201; source: Hunan China), Atractylodis Macrocephalae Rhizoma (batch number: 13,102,101; source: Zhejiang China) and Glycyrrhizae Radix et Rhizoma Praeparata Cum Melle (batch number: 13,112,501; source: Gansu China) was made from Guoda pharmacy (Shenyang, China), which followed identification by Professor Jingming Jia (Department of TCM, Shenyang Pharmaceutical University, Shenyang, China). The key source of the reference compounds (purity > 98%), including atractyloside A, schaftoside, ononin and glycyrrhizic acid was Shanghai yuanye Bio-Technology Co., Ltd. (Shanghai, China); ginsenoside Re, ginsenoside Rb1, formononetin, atractylenolide III and atractylenolide II were attained from the National Institute for the Control of Pharmaceutical and Biological Products (Beijing, China). In addition to that, Acetonitrile of HPLC grade, together with formic acid of LC-MS grade was attained from Fisher Scientific (Fair Lawn, NJ, USA), followed by attaining the purified water from Wahaha (Hangzhou, China).
The use of an Agilent 1260 UHPLC system (USA) was made in order to perform the chromatographic analysis. The use of a Universal XB C8 column (150 mm × 2.1 mm, 1.8 μm; Kromat, USA) having the column temperature of 45 °C was made. The mobile phase comprised 0.2% formic acid (A) and acetonitrile (B), which was conducted with gradient condition as hereunder: 2–2% (B) in 0 to 3 min, 2–22% (B) in 3 to 13 min, 22–70% (B) in 13 to 45 min. The setting of the flow rate at 0.20 mL·min−1 was ensured, together the injection volume amounting to be 2 μL. The use of a Bruker Solarix7.0 T FT-ICR-MS system (Bruker, Germany) as well as a Bruker Compass-Hystar workstation (Bruker, Germany) was made to carry out the mass spectra analysis. Ionization was carried out using positive as well as negative electrospray ionization (ESI) modes, followed by setting the optimized conditions as hereunder: nebulizer gas pressure, 4.0 bar; dry gas flow rate, 8 L·min−1; dry gas temperature, 200 °C; ion accumulation time, 0.15 s; time of flight, 0.6 ms; capillary voltage, 4.5 kV; and end plate offset, 500 V. Recording of the Full-scan mass spectrum data was performed between m/z 100 and 3000 amu. In respect of auto MS/MS mode, selection of both MS/MS Boost and MS/MS Isolation was made; moreover, range of the collision power lied between 10 eV and 30 eV as regards the MS/MS experimentations.
2.2. Preparation of SJZD for analysis
3. Results and discussion
In accordance with the documentary records of SJZD, four key constituting herbs that included Radix Ginseng (100 g), Poria (100 g), Rhizoma Atractylodis Macrocephalae (100 g), and Glycyrrhizae Radix et Rhizoma Praeparata Cum Melle (50 g) were crushed and made to form small pieces, followed by mixing and decocting two times in 3500 mL water for a period of 1 h each time in a glass flask [2,6]. Subsequently, mixing of the ensuing solution was done that followed drying with the help of lyophilization. Prior to the analysis, dissolution
The Fig. 1 reveals the base peak ion chromatograms (BPC) of SJZD in positive as well as negative ion modes, together with the respective compounds. The extract ion chromatograms (EIC) of each molecule weight were correspondingly attained for detecting the associated compound followed by presenting in Supporting information Fig. S1. Among the compounds identified, accurate identification of nine was carried out with the help of comparison with the retention time (tR) as well as the MS/MS data associated with the reference compounds in
2.3. Instrument and analytical conditions
2.1. Chemicals and materials
Fig. 1. The base peak ion chromatograms (BPC) of SJZD in both positive (A) and negative (B) ion modes and the corresponding compounds (C). 1-Atractyloside A, 2schaftoside, 3-ononin, 4-ginsenoside Re, 5-ginsenoside Rb1, 6-formononetin, 7-glycyrrhizic acid, 8-atractylenolide II, 9-atractylenolide III.
12
Journal of Chromatography B 1086 (2018) 11–22
Z. Guan et al.
Fig. 2. The chemical structures of chemical compounds detected in Radix Ginseng.
positive ion mode (Fig. 1). Moreover, the others were inferred by the retention times, in addition to molecular weight as well as MS/MS fragments. The use of the Bruker workstation was made for computing the molecular formulas of compounds through comparison with the identified molecular weights with the measured molecular weights, followed by accepting the error values being < 5.0 ppm. Further speculations of the layouts of the compounds were made using the MS/MS data. In aggregate, identification and characterization of 120 compounds were performed, whereas their layouts were presented in the Figs. 2-6. The presentation of MS/MS spectra of the usual compounds was provided in the Fig. 7, together with displaying their possible fragmentation pathways in the Fig. 8. Inferences of each and every ingredient were carried out with the help of the molecule formula and fragmentation pathways, followed by additional confirmation by reference to pertinent literature [7–16]. The Table 1 sheds light on the retention time, formula, molecular weight, calculated m/z, detected m/ z, error value and MS/MS data of the ingredients. Concrete illustration of the characterization of the ingredients in the individual lab materials was done as hereunder. 3.1. Characterization of constituents in Radix ginseng Saponins made the key active ingredients in Radix Ginseng, whereby, the majority of them were the glycosides of triterpenoid aglycones. In this study, tentative characterization of 34 triterpenoid saponins of Radix Ginseng in SJZD was carried out, followed by identifying 2 of them. Both Peak 4 and 5 in the Fig. 1C were ginsenoside Re as well as ginsenoside Rb1, correspondingly. Ginsenoside Rs5 is termed as a common ginsenoside, being less often stated in the literature, was put to use as an illustration for demonstrating the fragmentation pathways (Figs. 7A, 8A). In respect of the positive mode, the ion at m/z 831.48661 was deducted for being the adduct ion ([M + Na]+) and calculation of the formula was made as C44H72O13. The key fragment ions found in the MS/MS spectrum were at the m/z735.64263, 407.35689, 279.11181 as well as 181.07431. The ion at m/z 735.64263 revealed the loss of C2H2O3 (74 Da) from the precursor ion, whereas, the ion at m/z 407.35689 denoted the loss of C12H28O10 (328 Da) from m/z 735.64263. The ions at m/z 279.11181 as well as 181.07431 represented C11H18O8 and glucose, correspondingly. The characterization of other ingredient in Radix Ginseng was performed based on the fragmentation patterns and related literatures [7–10]. 3.2. Characterization of constituents in Poria 16 triterpene carboxylic acids made the key active ingredients in Poria [17]. In this research work, tentative characterization of 15 sixteen triterpene carboxylic acids of Poria in SJZD was performed. The use of Trametenolic acid, being a common composition of Poria was made as an instance for demonstrating the fragmentation pathways (Figs. 7B, 8B). In respect of the positive mode, inference of the ion at m/ z 421.33572 was made for being the key fragment ion, followed by indicating the loss of H2O2 (36 Da) from the precursor ion. The ion at m/z 2851.17113 in the MS/MS spectrum was an indication of the loss of C14H24 (192 Da) from the precursor ion. Characterization of the other constituents in Poria was carried out based on the fragmentation patterns and related literatures [11,12]. 3.3. Characterization of constituents in Rhizoma Atractylodis Macrocephalae Sesquiterpenes made the key active constitutes in Rhizoma Atractylodis Macrocephalae [18]. In this research work, tentative 13
Journal of Chromatography B 1086 (2018) 11–22
Z. Guan et al.
Fig. 3. The chemical structures of chemical compounds detected in Poria.
on the fragmentation patterns and related literatures [13,14].
characterization of 11 sesquiterpenes of Rhizoma Atractylodis Macrocephalae in SJZD was carried out, followed by precisely identifying three of them. Peak 1, 8 and 9 in the Fig. 1C were atractyloside A, atractylenolide II and atractylenolide, correspondingly. The use of Atractylenolide I, being a common composition of Rhizoma Atractylodis Macrocephalae was made as an instance for demonstrating the fragmentation pathways (Figs. 7C, 8C). In respect of the positive mode, inference of the ion at m/z 233.15386 was performed for being the adduct ion ([M + H]+), followed by calculation of the formula as C15H20O2. The key fragment ions found in the MS/MS spectrum were at the m/z 215.14072, 187.14559, 177.08796 as well as 145.10055. The ion at m/z 215.14072 was an indication of the loss of H2O (18 Da) from the precursor ion. The ions at m/z 187.14559 as well as 177.08796 were the indications of the loss of CO (28 Da) and C3H2 (38 Da) from m/ z 215.14072. Moreover, the ions at m/z 145.10055 suggested the loss of CH4O (32 Da) from m/z 177.08796. Characterization of the other ingredients in Rhizoma Atractylodis Macrocephalae was performed based
3.4. Characterization of constituents in Glycyrrhizae Radix et Rhizoma Praeparata Cum Melle Triterpenesaponins, together with several kinds of flavonoids, made the key active constituents in Glycyrrhizae Radix et Rhizoma Praeparata Cum Melle [15]. In this research work, tentative characterization of 60 ingredients of Glycyrrhizae Radix et Rhizoma Praeparata Cum Melle in SJZD was performed, followed by precise identification of 4 among them. Peak 2, 3, 6 and 7 presented in the in Fig. 1C represented schaftoside, ononin, formononetin and glycyrrhizic acid, correspondingly. The use of Liquiritigenin, being a common flavonoids composition of Glycyrrhizae Radix et Rhizoma Praeparata Cum Melle was made as an illustration for demonstrating the fragmentation pathways (Fig. 7D, 8D). In respect of the positive mode, inference of the ion at m/z 257.08089 was performed for being the adduct ion
14
Journal of Chromatography B 1086 (2018) 11–22
Z. Guan et al.
Fig. 4. The chemical structures of chemical compounds detected in Rhizoma Atractylodis Macrocephalae.
chemical ingredients existing in SJZD. The findings revealed the fact that ginsenosides (in Radix Ginseng), sixteen triterpene carboxylic acids (in Poria), Sesquiterpenes (in Rhizoma Atractylodis Macrocephalae), triterpenesaponins (in Glycyrrhizae Radix et Rhizoma Praeparata Cum Melle) as well as flavonoids (in Glycyrrhizae Radix et Rhizoma Praeparata Cum Melle) made the key ingredients existing in SJZD, followed by the provision of the particular data. This research provided more information of chemical compounds in SJZD than previous research in this field. Therefore, good amount of substantial foundation, together with considerable amount of information has been provided by this research work in respect of the pharmacological research of SJZD. This research together with other similar papers could be scientific proof for the chemical ingredients characterization on this method of other TCMFs. However, in this research, the chemical structures of compounds were speculated based on the precise molecule formula of MS and MS/ MS information and fragmentation pathways, followed by additional confirmation by reference to pertinent literature. The identification of structure was limited to known compounds only, which is also the limitation of chemical structure identification by mass spectrometry. Thus, the advantages of mass spectrometry identification were rapid and effective, for the more precise identification, assistance in other technologies such as nuclear magnetic resonance (NMR) and single crystal X-ray diffraction (SXRD) were required. Supplementary data to this article can be found online at https:// doi.org/10.1016/j.jchromb.2018.04.009.
([M + H]+), followed by the calculation of the formula as C15H13O4. The key fragment ions found in the MS/MS spectrum were at the m/z 147.04216, 137.02175 as well as 119.04821. The ion at m/z 147.04216 as well as137.02175 suggested the loss of C6H6O2 (110 Da) and C8H8O (120 Da) from the precursor ion, correspondingly. The ion at m/z 119.04821 denotes the loss of CO (28 Da) from m/z 147.04216. Uralsaponin N was used as a common triterpenesaponins composition of Glycyrrhizae Radix et Rhizoma Praeparata Cum Melle as an illustration for demonstrating the fragmentation pathways (Figs. 7E, 8E). In the positive mode, the ion at m/z 839.40598 was inferenced to be the adduct ion ([M + H]+), followed by the calculation of the formula as C42H63O17. The key fragment ions found in the MS/MS spectrum were at the m/z 663.37644, 487.32988 as well as 179.05516. The ion at m/z 663.37644 suggested the loss of C6H9O6 (178 Da) from the precursor ion which indicated the breaking of glycosidic bond. The ion at m/z 179.05516 was calculated as C6H10O6 as another fragment ion produced by the breaking of glycosidic bond. The ion at m/z 487.32988 suggested the loss of C6H9O6 (178 Da) from the precursor ion m/z 663.37644. Characterization of the other ingredients in Glycyrrhizae Radix et Rhizoma was performed based on the fragmentation patterns and related literatures [15,16]. 4. Conclusions A swift and straightforward methodology was performed to systematically characterize the chemical constituents found in SJZD with the help of UHPLC-FT-ICR-MS. This methodology brought forth shorttime as well as effective separation, together with detection of the
15
Journal of Chromatography B 1086 (2018) 11–22
Z. Guan et al.
Fig. 5. The chemical structures of flavonoids detected in Glycyrrhizae Radix et Rhizoma Praeparata Cum Melle.
16
Journal of Chromatography B 1086 (2018) 11–22
Z. Guan et al.
Fig. 6. The chemical structures of triterpenesaponins detected in Glycyrrhizae Radix et Rhizoma Praeparata Cum Melle.
17
Journal of Chromatography B 1086 (2018) 11–22
Z. Guan et al.
Fig. 7. The MS/MS spectra of the typical compounds. A-Ginsenoside Rs5, B-trametenolic acid, C-atractylenolide I, D-liquiritigenin, E-uralsaponin N.
18
Journal of Chromatography B 1086 (2018) 11–22
Z. Guan et al.
Fig. 8. The possible fragmentation pathways of the typical compounds. A-Ginsenoside Rs5, B-trametenolic acid, C-atractylenolide I, D-liquiritigenin, E-uralsaponin N.
19
Rt (min)
35.77 36.92 29.67 28.55 40.61 29.63 24.65 33.41 19.88 19.84 28.42 25.39 21.60 32.11 29.27 33.76 31.81 30.94 24.73 23.57 26.71 22.56 25.39 28.25 31.09 27.16 21.62 41.50 28.05 24.43 28.29 32.36 30.79 28.57 31.34
26.27 26.27 28.17 26.97 30.40 30.65
30.16
34.11
20.21 37.51 28.53 19.98 13.97 21.32
No.
1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35
36 37 38 39 40 41
42
43
44 45 46 47 48 49
Dehydroprotopanaxadiol II Dehydroprotopanaxatriol I Dehydroprotopanaxatriol II Panaxadione Ginsenoside Rk2 Ginsenoside Rk3 Ginsenoside Rh4 Compound K Ginsenoside F1 Ginsenoside Rh1 Ginsenoside ST2 Ocotillol derivative 3a Ginsenoside Km Ginsenoside Rs6 Compound Y Ginsenoside Rg6 Ginsenoside Rk1 Ginsenoside Rz1 Ginsenoside Rg5 Ginsenoside Rg3 Ginsenoside SL2 12,23-Epoxyginsenoside Rg1 Ginsenoside Rf Ginsenoside ORh1 Chikusetsusaponin IVa Floralginsenoside F Floralginsenoside E Ginsenoside Rs5 C-Y2 Ginsenoside Rg1 Ginsenoside Re Ginsenoside Ro Floralginsenoside Kb Ginsenoside Rb1 3β,16α-Dihydroxy-lanostane −7.9(11),24-Trien-21-oic acid Dehydrotrametenolic acid 16α-Hydroxytrametenolic acid Trametenolic acid Poricoic acid B 16-Deoxyporicoic acid B 3β-Hydroxy-16α-acetyloxy-lanosta −7.9(11),24-trien-21-oic acid 3-O-acetyl-16α-hydroxy -dehydrotrametenolic acid 3-O-acetyl-16α-hydroxytrametenolic acid Me dehydroeburicoate 6α-Hydroxydehydropachymic acid Poricoic acid DM Poricoic acid A Ganoderic acid B 3β,16α-Diacetoxy-24-(hydroxymethyl) lanost-8-en-21-oic acid
Identification
Monoisotopic mass (m/z) 443.38836 459.38327 459.38327 511.33940 605.44118 621.43797 621.43797 623.45175 639.44666 639.44666 655.44157 655.44157 655.44157 663.44666 755.49400 767.49400 767.49400 767.49400 767.49400 785.50457 799.48383 799.48383 801.49948 803.50701 817.43448 817.49440 817.49440 831.48651 845.49041 845.49041 947.55739 957.50536 965.49628 1153.60114 453.33632 455.35197 455.35197 457.36762 467.31559 469.33124 495.34689 495.34689 515.37310 521.33915 541.35346 529.35237 537.29768 555.27186 589.40988
Formula C30H51O2 C30H51O3 C30H51O3 C30H48O5Na C36H61O7 C36H61O8 C36H61O8 C36H63O8 C36H63O9 C36H63O9 C36H63O10 C36H63O10 C36H63O10 C38H63O9 C41H71O12 C42H71O12 C42H71O12 C42H71O12 C42H71O12 C42H73O13 C42H71O14 C42H71O14 C42H73O14 C44H76O10K C42H66O14Na C42H73O15 C42H73O15 C44H72O13Na C43H73O16 C43H73O16 C43H83O18 C48H77O19 C46H77O21 C55H93O25 C30H45O3 C30H47O3 C30H47O3 C30H49O3 C30H43O4 C30H45O4 C32H47O4 C32H47O4 C32H51O5 C32H50O3K C33H49O6 C32H49O6 C31H46O5K C30H44O7K C35H57O7
Table 1 The results of chemical constituent characterization of SJZD by FT-ICR-MS.
20 521.33716 541.35556 529.35323 537.29581 555.27363 589.41266
515.37173
495.34708
455.35238 455.35238 457.36812 467.31580 469.33261 495.34754
443.38986 459.38375 459.38483 511.34040 605.44157 621.43768 621.43797 623.45176 639.44633 639.44632 655.44311 655.44151 655.44142 663.44666 755.49483 767.49463 767.49621 767.49531 767.49425 785.50339 799.48337 799.48303 801.49803 803.50802 817.43492 817.49388 817.49342 831.48661 845.48747 845.48718 947.56099 957.50368 965.49370 1153.59692 453.33710
Detected mass (m/z)
−0.40
[M − H2O + H]+
[M + K]+ [M − H]− [M + H]+ [M + K]+ [M + K]+ [M + H]+
[M + H]
3.82 −3.88 −1.64 3.48 −3.19 −4.72
2.66
395.36772, 437.33189, 421.33572, 409.28485, 397.31662, 437.33282,
−0.89 −0.89 −1.10 −0.46 −2.93 −1.32 [M + H]+ [M − H2O + H]+ [M + H]+ [M − H2O + H]+ [M + H]+ [M − H2O + H]+
+
423.36375, 409.38346, 423.36409, 443.38708, 443.38874, 459.38368, 409.38312, 461.39789, 603.41985, 603.42033, 493.38126, 493.38315, 506.39120, 409.38422, 461.39637, 603.42658, 443.38763, 443.38900, 587.43633, 749.48820, 491.36956, 475.37688, 621.43579, 477.38776, 631.38351, 493.39046, 493.39607, 735.64263, 475.38546, 475.38796, 621.42643, 631.38258, 447.35577, 783.48952, 435.31284,
−3.40 −1.05 −3.40 −1.96 −0.65 −2.54 −3.02 −0.03 0.52 0.53 −2.35 0.09 0.23 0.00 −1.09 −0.82 −2.87 −1.70 −0.31 1.50 0.57 1.01 1.81 −1.26 −0.55 0.63 1.19 −0.11 3.47 3.82 −3.80 1.75 2.68 3.66 −1.73 [M + H]+ [M + H]+ [M + H]+ [M + Na]+ [M + H]+ [M + H]+ [M + H]+ [M + H]+ [M + H]+ [M + H]+ [M + H]+ [M + H]+ [M + H]+ [M + H]+ [M + H]+ [M + H]+ [M + H]+ [M + H]+ [M + H]+ [M + H]+ [M + H]+ [M + H]+ [M + H]+ [M + K]+ [M + Na]+ [M + H]+ [M + H]+ [M + Na]+ [M + HCOOH-H]− [M + HCOOH-H]− [M + H]+ [M + H]+ [M + HCOOH-H]− [M + HCOOH-H]− [M − H2O + H]+ 299.26112 297.25031 215.17499 277.20385 283.23173 297.25884
109.10256 69.07763 109.10193 69.07015 109.10267 109.10304 69.07651 165.06845 163.05846 163.05694 163.05773 163.06004 163.06129 69.07641 296.09754 147.06394 109.10288 163.05773 407.36994 441.3765 310.11906 165.06757 163.05988 290.16350 163.06396 163.05481 326.11348 287.06043 308.11691 179.05378 179.05255 177.03002 179.05482 341.10843 309.25784
467.38808, 501.35669, 441.29522, 426.31107, 481.28831, 505.38933,
409.37226, 447.31274, 397.30047, 381.30452, 325.20193, 425.37127,
P P P P P P
P
P
P P P P P P
G G G G G G G G G G G G G G G G G G G G G G G G G G G G G G G G G G P
Source
(continued on next page)
299.26462 403.32992 279.20338 279.20475 297.25088 297.25650
473.35293, 437.33901, 393.34220
435.31290, 391.32978, 295.23088
325.28528, 393.34681, 251.17113, 365.29466, 353.32037, 393.34113,
407.36852, 121.10114, 407.36769, 359.29484, 407.36706, 407.36829, 121.10288, 429.40112, 457.36497, 565.37326, 428.40582, 459.38149, 443.41207, 121.09831, 429.40128, 459.38674, 407.36596, 411.39626, 425.37169, 605.44168, 443.71847, 445.39620, 457.36484, 429.40254, 455.35669, 444.73498, 444.72972, 407.35689, 427.40177, 411.40398, 441.36383, 455.35381, 341.03776, 603.42612, 391.32005,
MS/MS (m/z)
ppm
Ion mode
Z. Guan et al.
Journal of Chromatography B 1086 (2018) 11–22
Rt (min)
34.66 21.04 37.90 34.66 23.87 32.63 42.52 24.83 16.74 15.50 14.91 25.26 20.81 24.43 31.98 19.19 22.84 24.52 32.26 36.13 35.38 33.28 35.44 34.26 2.65 33.86 31.98 29.58 39.17 37.83 35.71 27.64 18.16
27.39 35.71 18.08 20.58 17.14 31.38 24.43 20.30 22.75 17.38 20.30 41.52 20.22 20.60 23.64 18.68 18.98 16.14 19.88 17.64
No.
50 51 52 53 54 55 56 57 58 59 60 61 62 63 64 65 66 67 68 69 70 71 72 73 74 75 76 77 78 79 80 81 82
83 84 85 86 87 88 89 90 91 92 93 94 95 96 97 98 99 100 101 102
Table 1 (continued)
Atractylenolide II Atractylentrid Atractylenolide I Atractylenolide III Chrysin Selina-4(14),7(11)-dien-8-one Dibutyl phthalate Luteolin Icariside D1 Atractyloside A Atractyloside B Pinocembrin Liquiritigenin Formononetin Echinatin Vestitol Naringenin Licochalcone B Glabrene Glabrone Licochalcone A Licoflavonol 3′-O-methyl glabridin 4′-O-methyl glabridin Kumatakenin Glicoricone Licoarylcoumarin Glycycoumarin Glyasperins D Glycyrin Licoricone Licopyranocoumarin 7-Hydroxy-2-(4-hydroxyphenyl)-6,8-bis (3-methyl-2-buten-1-yl)-2,3-dihydro-4Hchromen-4-one Kanzonol M Kanzonol O 6,8-Diprenylgenistein Isoliquiritin Liquiritin Kanzonol F Ononin Vitexin Choerospondin Licoagroside B Trifolirhizin Glycyrrhetinic acid Licoagroside A liquiritin Apioside Isoliquiritin apioside Schaftoside Isoschaftoside Violanthin Isoviolanthin Vicenin-2
Identification
21 399.18022 405.13086 407.18530 419.13366 419.13366 421.20095 431.13366 433.11292 433.11402 433.13405 447.12857 471.34689 491.11950 551.17592 551.17592 565.15518 565.15518 579.17083 579.17083 595.16575
231.13796 233.11722 233.15361 249.14852 253.05063 257.13022 279.15909 285.04046 439.15747 471.22007 495.24470 257.08084 257.08084 269.08084 271.09649 271.11214 273.07575 285.07685 321.11323 335.09250 337.14453 355.11761 355.15400 355.15400 359.07724 369.13326 369.13326 369.13326 371.18530 383.14891 383.14891 385.12818 393.20604
C15H19O2 C14H17O3 C15H21O2 C15H21O3 C15H9O4 C15H22OK C16H23O4 C15H9O6 C19H28O10Na C21H36O10Na C22H39O12 C15H13O4 C15H13O4 C16H13O4 C16H15O4 C16H17O4 C15H13O5 C16H13O5 C20H17O4 C20H15O5 C21H21O4 C20H19O6 C21H23O5 C21H23O5 C18H15O8 C21H21O6 C21H21O6 C21H21O6 C22H26O5 C22H23O6 C22H23O6 C21H21O7 C25H29O4
C23H27O6 C22H22O6Na C25H27O5 C21H23O9 C21H23O9 C26H29O5 C22H23O9 C21H21O10 C21H21O10 C18H25O12 C22H23O10 C30H47O4 C23H23O12 C26H31O13 C26H31O13 C26H29O14 C26H29O14 C27H31O14 C27H31O14 C27H31O15
Monoisotopic mass (m/z)
Formula
399.17838 405.13141 407.18672 419.13391 419.13375 421.20184 431.13320 433.11297 433.11249 433.13396 447.12865 471.34787 491.11738 551.17559 551.17577 565.15456 565.15500 579.17010 579.17031 595.16480
231.13815 233.11680 233.15386 249.14875 253.05018 257.13052 279.15956 285.03959 439.15746 471.21966 495.24294 257.08103 257.08089 269.08117 271.09637 273.11218 273.07594 285.07627 321.11278 335.09185 337.14387 355.11809 355.15440 355.15431 359.07639 369.13363 369.13357 369.13426 371.18615 383.14945 383.14927 385.12846 393.20743
Detected mass (m/z)
4.61 −1.35 −3.49 −0.60 −0.22 −2.11 1.06 −0.11 3.52 0.21 −0.16 −2.09 4.32 0.59 0.27 1.10 0.33 1.27 0.90 1.60
[M + H]+ [M + Na]+ [M + H]+ [M + H]+ [M + H]+ [M + H]+ [M + H]+ [M + H]+ [M − H]− [M + H]+ [M + H]+ [M + H]+ [M − H]− [M + H]+ [M + H]+ [M + H]+ [M + H]+ [M + H]+ [M + H]+ [M + H]+
367.15793, 375.13794, 337.11593, 309.10743, 282.11531, 389.17549, 399.11572, 269.04531, 282.11643, 309.10225, 285.07128, 339.26538, 327.07693, 417.12641, 417.12548, 446.11564, 446.11986, 460.12586, 460.12536, 476.12561,
213.12351, 185.13547, 215.14072, 231.13685, 209.06399, 202.72643, 205.08378, 201.01564, 285.13978, 287.18664, 287.19446, 152.01593, 147.04216, 237.06352, 239.07549, 241.09571, 152.01597, 255.07891, 270.09511, 281.06382, 307.13591, 299.06137, 323.13596, 307.13521, 345.06571, 313.07536, 353.10593, 337.11578, 339.16597, 351.12591, 327.09253, 353.10521, 323.13571,
−0.86 1.80 −1.10 −0.92 1.79 −1.16 −1.69 3.07 0.01 0.87 3.55 −0.75 −0.23 −1.26 0.44 −0.15 −0.70 2.04 1.40 1.92 1.97 −1.34 −1.13 −0.88 2.37 −0.98 −0.84 −2.69 −2.30 −1.40 −0.93 −0.73 −3.53 [M + H] [M + H]+ [M + H]+ [M + H]+ [M − H]− [M + K]+ [M + H]+ [M − H]− [M + Na]+ [M + Na]+ [M + HCOOH-H]− [M + H]+ [M + H]+ [M + H]+ [M + H]+ [M + H]+ [M + H]+ [M − H]− [M − H]− [M − H]− [M − H]− [M + H]+ [M + H]+ [M + H]+ [M + HCOOH-H]− [M + H]+ [M + H]+ [M + H]+ [M + H]+ [M + H]+ [M + H]+ [M + H]+ [M + H]+
175.06651 83.04337 145.10055 187.14653 107.01556 105.06856 57.06455 107.01345 79.05661 139.14358 137.14063 104.06359 119.04821 136.02586 121.03782 123.04827 120.06783 165.06538 214.10529 136.02593 217.12435 165.02596 285.11495 269.12734 328.06134 136.02543 301.07541 206.09162 303.12483 315.09157 312.06586 162.03716 138.03632
Gly Gly Gly Gly Gly Gly Gly Gly Gly Gly Gly Gly Gly Gly Gly Gly Gly Gly Gly Gly
A A A A A A A A A A A Gly Gly Gly Gly Gly Gly Gly Gly Gly Gly Gly Gly Gly Gly Gly Gly Gly Gly Gly Gly Gly Gly
Source
(continued on next page)
338.15431, 331.12769 353.11682, 337.12583 268.04196, 155.03697 281.10593, 255.07495 255.07461, 136.02561 367.15319, 353.14128 267.07593, 122.04536 163.06321,154.03257 271.06593, 152.01479 147.05422, 127.02467 229.08474, 149.02177 189.16722, 137.13835 164.03532, 148.03643 255.07631, 136.02758 255.07953, 137.02497 431.10298, 401.09589 431.10358, 401.09658 431.10581, 415.10692 431.10857, 415.10486 431.10592, 270.05391
185.12567, 153.08055, 187.14559, 215.13995, 143.05648, 178.96543, 149.02367, 133.02614, 123.07536, 183.13114, 183.14758, 124.02671, 137.02175, 148.05293, 149.06349, 149.06493, 124.02641, 193.05761, 253.09137, 267.07936, 245.12961, 271.05284, 301.11679, 285.11357, 344.06257, 299.06351, 337.11593, 301.07152, 309.15674, 321.11539, 315.08961, 222.09143, 254.06395,
MS/MS (m/z)
+
ppm
Ion mode
Z. Guan et al.
Journal of Chromatography B 1086 (2018) 11–22
Journal of Chromatography B 1086 (2018) 11–22
3.18 −0.05 −0.58 0.85 0.93 −2.13 1.10 0.65 −0.11 [M − H]− [M + H]+ [M + Na]+ [M + H]+ [M + H]+ [M + Na]+ [M + Na]+ [M + H]+ [M + H]+ 849.35234 881.41659 887.40409 897.41069 955.48882 979.48939 979.48622 985.46325 1027.47457 849.35504 881.41654 887.40357 897.41146 955.48971 979.48730 979.48730 985.46389 1027.47445 27.85 28.47 30.67 27.37 26.97 29.83 32.38 27.84 27.92 112 113 114 115 116 117 118 119 120
Uralsaponin E 22β-Acetoxyl-glycyrrhizin 22β-Acetoxyl-glycyrrhaldehyde Uralsaponin F Uralsaponin T Yunganoside A1 Yunganoside B1 Licorice-saponin A3 Uralsaponin X
C42H57O18 C44H65O18 C44H64O17Na C44H65O19 C48H75O19 C48H76O19Na C48H76O19Na C48H73O21 C50H75O22
694.21268 697.21272 807.41569 821.39689 823.41322 825.42596 831.41360 839.40690 694.21303 697.21270 807.41615 821.39541 823.41106 825.42671 831.41374 839.40598 C35H36O14N C35H37O15 C42H63O15 C42H61O16 C42H63O16 C42H65O16 C42H64O15Na C42H63O17 Licorice glycoside E Licorice glycoside B Uralsaponin W Uralsaponin O Glycyrrhizic acid Uralsaponin P Licorice-saponin B2 Uralsaponin N 26.03 24.48 34.83 30.35 31.38 26.29 32.53 30.47 103 104 105 106 107 108 109 111
“The authors declare that there are no conflicts of interest”. Acknowledgements The authors gratefully acknowledge the facilities and assistance provided by Dr. Fei Han of the Bruker Corporation. G represented Ginseng Radix et Rhizoma; P represented Poria; A represented Atractylodis Macrocephalae Rhizoma; Gly represented Glycyrrhizae Radix et Rhizoma Praeparata Cum Melle.
Gly Gly Gly Gly Gly Gly Gly Gly Gly 179.04641 179.04753 179.04966 177.06586 179.04616 149.07233 149.07347 179.04862 149.08465 485.31611, 529.34658, 513.34358, 545.33421, 458.35522, 459.37546, 459.37235, 472.33458, 529.34414,
Gly Gly Gly Gly Gly Gly Gly Gly 294.08615, 257.07166 297.08341, 257.07246 475.37620, 181.06385 491.36486, 181.06344 471.32977, 179.04623 487.33574, 165.06255 238.17058, 224.19570 487. 32,988.
440.14134, 443.14235, 653.41902, 669.41376, 647.36821, 663.36548, 413.37584, 663.37644, 179.05516 661.34962, 705.37623, 689.37723, 721.36447, 779.44623, 811.43655, 811.43913, 809.42963, 705.37552, 0.51 −0.04 0.56 −1.80 −2.74 0.91 0.17 −1.10 [M + H] [M + H]+ [M + H]+ [M + H]+ [M + H]+ [M + H]+ [M + Na]+ [M + H]+
MS/MS (m/z)
+
Ion mode Detected mass (m/z) Monoisotopic mass (m/z) Formula Identification Rt (min) No.
Table 1 (continued)
Conflicts of interest
ppm
Source
Z. Guan et al.
Funding This work was supported by the National Natural Science Foundation of China (NSFC: 81503035). This work was supported by the Shenyang Pharmaceutical University Young Teachers' Career Development Program. References [1] Kang An, Guo Jin-rui, Zhang Zhen, Wang Xiao-long, Simultaneous quantification of ten active components in traditional Chinese formula Sijunzi decoction using a UPLC-PDA method, J. Anal. Methods Chem. 570359 (2014) 2–8. [2] Yang Liu, Junshan Yang, Zongwei Cai, Chemical investigation on Sijunzi decoction and its two major herbs Panax ginseng and Glycyrrhiza uralensis by LC/MS/MS, J. Pharm. Biomed. Sci. 41 (2006) 1642−1647. [3] Zhibo Guan, Juan Wu, Cancan Wang, Fang Zhang, Yinan Wang, Miao Wang, Min Zhao, Chunjie Zhao, Investigation of the preventive effect of Sijunzi decoction on mitomycin C induced immunotoxicity in rats by 1H NMR and MS-based untargeted metabolomic analysis, J. Ethnopharmacol. 210 (2018) 179–191. [4] Yinan Wang, Fei Han, Aihua Song, Miao Wang, Min Zhao, Chunjie Zhao, Rapid characterization of the chemical constituents of Cortex Fraxini by homogenate extraction followed by UHPLC coupled with Fourier transform ion cyclotron resonance mass spectrometry and GC–MS, J. Sep. Sci. 00 (2016) 1–10. [5] Ting Liu, Xiumin Tian, Zhaoqin Li, Fei Han, Bin Ji, Yunli Zhao, Zhiguo Yu, Metabolic profiling of Gegenqinlian decoction in rat plasma, urine, bile and feces after oral administration by ultra high performance liquid chromatography coupled with Fourier transform ion cyclotron resonance mass spectrometry, J. Chromatogr. B 1079 (2018) 69–84. [6] Qiuxiang Xu, Hongli Yu, Hao Wu, Fenqiang You, Yunhan Zhu, Lijuan Tang, Pharmacological comparison between traditional decoction and machine decoction of Sijunzi Tang, Chin. J. Exp. Tradit. Med. Formulae. 19 (2013) 250–255. [7] Byong-Kyu Shin, Sung Won Kwon, Jeong Hill Park, Chemical diversity of ginseng saponins from Panax ginseng, J. Ginseng Res. 39 (2015) 287−298. [8] Zai-Qun Liu, Chemical insights into ginseng as a resource for natural antioxidants, Chem. Rev. 112 (2012) 3329−3355. [9] JaeWon Lee, Ram Choi, Young-Chang Kim, Doo Jin Choi, Young-Seob Lee, GeumSoog Kim, Nam-In Baek, Seung-Yu Kim, Dae Young Lee, Comprehensive profiling and quantification of ginsenosides in the root, stem, leaf, and berry of Panax ginseng by UPLC-QTOF/MS, Molecules 22 (2017) 2147. [10] Lei Zhang, Qi-Le Zhou, Xiu-Wei Yang, Determination of the transformation of ginsenosides in Ginseng Radix et Rhizoma during decoction with water using ultrafast liquid chromatography coupled with tandem mass spectrometry, J. Sep. Sci. 41 (2018) 1039–1049. [11] Bing Xia, Yan Zhou, Hong Sheng Tan, Sheng Ding Li, Xu Hong Xi, Advanced ultraperformance liquid chromatography–photodiode array–quadrupole time-of-flight mass spectrometric methods for simultaneous screening and quantification of triterpenoids in Poria cocos, Food Chem. 152 (2014) 237–244. [12] José-Luis Ríos, Chemical constituents and pharmacological properties of Poria cocos, Planta Med. 77 (2011) 681–691. [13] Guo-Shun Shan, Liang-Xiao Zhang, Qi-Miao Zhao, Hong-Bin Xiao, Rong-Jie Zhuo, Gang Xua, Hong Jiang, Xian-Min You, Tian-Zhu Jia, Metabolomic study of raw and processed Atractylodes macrocephala Koidz by LC–MS, J. Pharm. Biomed. Anal. 98 (2014) 74–84. [14] Xu Yangyang, Hao Cai, Gang Cao, Yu Duan, Pei Ke, Tu Sicong, Jia Zhou, Xie Li, Dongdong Sun, Jiayu Zhao, Jing Liu, Xiaoqi Wang, Lin Shen, Profiling and analysis of multiple constituents in Baizhu Shaoyao San before and after processing by stirfrying using UHPLC/Q-TOF-MS/MS coupled with multivariate statistical analysis, J. Chromatogr. B 1083 (2018) 110–123. [15] Ci-Hai Zhang, Yue Yu, Yi-Zeng Liang, Xiao-Qing Chen, Purification, partial characterization and antioxidant activity ofpolysaccharides from Glycyrrhiza uralensis, Int. J. Biol. Macromol. 79 (2015) 681–686. [16] Wei Song, Longlong Si, Shuai Ji, Han Wang, Xiao-mei Fang, Yu Li-yan, Ren-yong Li, Li-na Liang, Demin Zhou, Min Ye, Uralsaponins M−Y, antiviral triterpenoid saponins from the roots of Glycyrrhiza uralensis, J. Nat. Prod. 77 (2014) 1632–1643. [17] Haruo Nukaya, Hirokazu Yamashiro, Hirotatsu Fukazawa, Hitoshi Ishida, Kuniro Tsuji, Isolation of inhibitors of TPA-induced mouse ear edema from Hoelen, Poria cocos, Chem. Pharm. Bull. 44 (1996) 847–849. [18] Qinhua Chen, Hongsheng He, Peng Li, Jun Zhu, Min Xiong, Identification and quantification of atractylenolide I and atractylenolide III in Rhizoma Atractylodes Macrocephala by liquid chromatography–ion trap mass spectrometry, Biomed. Chromatogr. 27 (2013) 699–707.
22