High-resolution Tandem Mass Spectrometry for Studying Chemical Constituents of Gynura bicolor DC
Özet
We describe a general protocol that integrates high-resolution mass spectrometry analysis and molecular docking in traditional Chinese medicine research.
Abstract
The separation and analysis of the desired chemical components are important subjects for the fundamental research of traditional Chinese medicine (TCM). Ultra-high-performance liquid chromatography quadrupole time-of-flight tandem mass spectrometry (UPLC-Q-TOF-MS/MS) has gradually become a leading technology for the identification of TCM ingredients. Gynura bicolor DC. (BFH), a perennial stemless herb used for medicine and food in China has medicinal effects such as clearing heat, moistening the lung, relieving cough, dispersing stasis, and relieving swelling. Polyphenols and flavonoids contain numerous isomers, which hinder the identification of the complex compounds in BFH. This paper presents a systematic protocol for studying chemical constituents of BFH based on solvent extraction and integrated data via UPLC-Q-TOF-MS.
The method described here includes systematic protocols for sample pretreatment, MS calibration, MS acquisition, data processing, and analysis of results. Sample pretreatment includes collection, cleaning, drying, crushing, and extraction. MS calibration consists of multipoint and single-point correction. Data processing includes data importing, method establishment, analysis processing, and result presentation. Representative results of the typical fragmentation pattern of phenolic acids, esters, and glycosides in Gynura bicolor DC. (BFH) are presented in this paper. In addition, organic solvent selection, extraction, data integration, collision energy selection, and method improvement are discussed in detail. This universal protocol can be widely used to identify complex compounds in TCM.
Introduction
Traditional Chinese medicine (TCM) has been clinically practiced in China for thousands of years, and it plays a vital role in maintaining the health of Chinese people1. The composition of TCM is diverse and complex, and TCM has been widely reported in many qualitative studies focusing on the chemical composition2. The chemical components in TCM can be roughly divided into the following categories such as alkaloids, organic acids, phenylpropanoids, coumarins, lignans, quinones, flavonoids, terpenoids, triterpenoid saponins, steroid saponins, cardiac glycosides, and tannins3. Given the large numbers of unknown components and indistinguishable isomers in TCM, the separation and analysis of the desired chemical components are important subjects for the fundamental research of TCM4.
Ultra-high-performance liquid chromatography quadrupole time-of-flight mass spectrometry (UPLC-Q-TOF-MS) has been applied to analyze substances in traditional Chinese medicine (TCM), which can be separated by ultra-high-performance liquid chromatography5,6. The high resolution of MS can provide extensive ion information, which is used for database analysis with error less than 5 ppm7. After turning on the collision energy, the secondary MS mode can obtain secondary fragment ions, whose intensity and number are affected by the magnitude of energy8.
Gynura bicolor DC. (BFH), a perennial stemless herb widely used for medicine and food (Figure 1A), is a rare and endangered plant unique to China9. BFH has abundant anthocyanins, polyphenols, flavonoids, and strong antioxidant capacity10. BFH has medicinal effects including clearing heat, cooling blood, moistening the lung, relieving cough, dispersing stasis, relieving swelling, relieving summer heat, and eliminating heat. Few studies have focused on the chemical composition of BFH11. Polyphenols and flavonoids contain numerous isomers, which make the identification of the complex compounds in BFH difficult. A universal method for the identification of chemical components must be developed, which can be applied to all kinds of TCM. This study aimed to report a systematic protocol for studying chemical constituents of BFH based on solvent extraction and integrated data via UPLC-Q-TOF-MS.
Protocol
Representative Results
Discussion
Besides water decoction12, organic solvent extraction is another common method of TCM pretreatment13. According to the principle of similar phase dissolution, numerous components have been extracted by the combination of various organic solvents14. Ultrasonic-assisted extraction is one of the main methods used to obtain components in TCM15. Supercritical carbon dioxide extraction is good for extracting certain kinds of substan…
Açıklamalar
The authors have nothing to disclose.
Acknowledgements
This work was funded by the National Natural Science Foundation of China (82104881), Inheritance and Innovation Team of TCM Treatment of Immune Diseases, Chongqing Medical Scientific Research Project (Joint project of Chongqing Health Commission and Science and Technology Bureau) (2022DBXM007), A special project for performance incentive and guidance of Chongqing Scientific Research Institute (cstc2022jxjl120005), A special project for Chongqing Postdoctoral Science Foundation (2022CQBSHTB3035), Senior Medical Talents Program of Chongqing for Yong and Middle-aged, the Program for Scientific Institutions of Chongqing (independent research project No.2022GDRC015).
Materials
| chloroform | Sinopharm Chemical ReagentCo., Ltd | CAS 67-66-3 | |
| ethanol | ChuandongChemical | CAS 64-17-5 | |
| ethyl acetate | ChuandongChemical | CAS 141-78-6 | |
| liquid chromatograph | Waters | ACQUITY Class 1 plus | |
| MassLynx | Waters | V4.2 | MS control software |
| n-butyl alcohol | ChuandongChemical | CAS 71-36-3 | |
| petroleum ether | ChuandongChemical | CAS 8032-32-4 | |
| Quadrupole time-of-flight mass spectrometry | Waters | SYNAPT XS | |
| UNIFI | Waters | Data analysis software |
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