CN-115932029-B - Mass spectrum imaging method for analyzing chemical components and spatial distribution of dried traditional Chinese medicine decoction pieces by adopting imprinting method

CN115932029BCN 115932029 BCN115932029 BCN 115932029BCN-115932029-B

Abstract

The invention provides a mass spectrum imaging method for analyzing chemical components and spatial distribution of dry traditional Chinese medicine decoction pieces by adopting a imprinting method, which comprises the steps of exerting pressure on the traditional Chinese medicine decoction pieces to be detected to obtain a relatively flat section, establishing a sandwich structure, transferring the section components of the traditional Chinese medicine decoction pieces to a transfer film by adopting the imprinting method, carrying out ionization scanning on the transfer film in a mass spectrometer by adopting a mass spectrum imaging detection method to obtain mass spectrum imaging data of the chemical components in the transfer film of the traditional Chinese medicine decoction pieces to be detected, carrying out data processing on the mass spectrum imaging data, identifying the chemical components in a sample to be detected, and analyzing the spatial distribution rule of metabolites in the traditional Chinese medicine decoction pieces to be detected. The method can overcome the difficulties of drying, hardness and uneven surface of the traditional Chinese medicine decoction pieces and difficult preparation of imaging samples, overcomes the defects of a mass spectrometry imaging method of the dried traditional Chinese medicine decoction pieces, is simple and convenient to operate, has high sensitivity, wide dynamic range and multiple component coverage types, and has remarkable advantages.

Inventors

LI MINGLI
LAI JUN
YANG TINGTING
HU ZHE
PENG ZHANGXIAO
FU YANLEI
SHU LIEBO

Assignees

上海鹿明生物科技有限公司

Dates

Publication Date: 20260508
Application Date: 20221207

Claims (9)

1. A mass spectrum imaging method for analyzing chemical components and spatial distribution of dried traditional Chinese medicine decoction pieces by adopting a blotting method is characterized by comprising the following steps: Step S1, applying pressure to the traditional Chinese medicine decoction pieces to be detected to obtain a relatively flat section; S2, establishing a sandwich structure, and transferring the components of the section of the Chinese medicinal decoction pieces to be tested onto a transfer film by adopting a blotting method; Spraying 300-500 mu L of 80% acetonitrile solution on a transfer film, placing the traditional Chinese medicine decoction pieces to be tested on the wetted transfer film, sequentially covering water absorbing paper, a rubber plate and sponge paper on two sides of a transfer film-traditional Chinese medicine decoction piece sample pair, applying pressure of 0.3-0.5 ton to the transfer film, applying pressure for 5-10S, and finally adhering the transfer film on a glass slide; S3, performing ionization scanning on the transfer film in a mass spectrometer by adopting a mass spectrometry imaging detection method to obtain mass spectrometry imaging data of chemical components of the traditional Chinese medicine decoction pieces to be detected on the transfer film; s4, performing data processing on the mass spectrum imaging data, and identifying chemical components in the traditional Chinese medicine decoction pieces to be detected; And S5, analyzing the spatial distribution rule of metabolites in the traditional Chinese medicine decoction pieces to be detected.
2. The mass spectrometry imaging method for analyzing chemical components and spatial distribution of dried traditional Chinese medicine decoction pieces by adopting a blotting method according to claim 1, wherein the step S1 is characterized in that the traditional Chinese medicine decoction pieces with relatively flat cross sections and thinner thickness are selected, and pressure of 0.3-0.5 ton is applied to the traditional Chinese medicine decoction pieces, and the pressure is applied for 5-10S, so that the traditional Chinese medicine decoction pieces with flat cross sections to be measured are obtained.
3. The mass spectrometry imaging method for analyzing chemical components and spatial distribution of dried decoction pieces of traditional Chinese medicine by blotting method according to claim 1, wherein in step S2, the transfer membrane is a nylon 66 microporous membrane.
4. The mass spectrometry imaging method for analyzing chemical components and spatial distribution of dried traditional Chinese medicine decoction pieces by adopting a blotting method according to claim 3, wherein in the step S2, 300-500 mu L of 80% acetonitrile solution is sprayed on the nylon 66 microporous filter membrane, so that the transfer membrane is wet and has no obvious water stain; and/or the number of the groups of groups, The diameter of the nylon 66 microporous filter membrane is 50 mm, and the pore diameter is 0.22 mu m.
5. The mass spectrometry imaging method for analyzing chemical components and spatial distribution of dried decoction pieces of traditional Chinese medicine by blotting according to claim 1, wherein in step S3, the mass spectrometer parameters are set as follows: The mass spectrometer is a Q-Orbitrap mass spectrometer, a adopted platform is a AFADESI-MSI platform, a spray solvent is an 80% acetonitrile solution, the solvent flow rate is 5 mu L/min, the air extraction flow rate is 45L/min, the distance between the spray and a slice is 0.5-0.8 mm, the distance between the surface of a sample and the sprayer is 2-4 mm, and the distance between the sprayer and an ion conveying pipe is 2-4 mm.
6. The mass spectrometry imaging method for analyzing chemical components and spatial distribution of dried decoction pieces of Chinese medicine by blotting according to claim 1, wherein in step S3, parameters of the ionization scan are set as follows: Positive ion Scan, full Scan mode, MS resolution of 70000, mass range of 70-1000 Da, automatic gain control target of 3E6, maximum injection time of 200 MS, capillary temperature of 350 ℃.
7. The mass spectrometry imaging method for analyzing chemical components and spatial distribution of dried decoction pieces of Chinese medicinal materials by blotting according to claim 1, wherein in step S4, the software for data processing comprises imzMLConverter and a Cardinal software package; and/or the number of the groups of groups, In step S4, the data preprocessing includes: and converting the original data into a imzML format file through imzMLConverter processing software, and importing the imzML format file into a Cardinal software package to perform background subtraction, peak alignment and peak screening processing to obtain a mass spectrum imaging diagram of the metabolite.
8. The mass spectrometry imaging method for analyzing chemical components and spatial distribution of dried traditional Chinese medicine decoction pieces by adopting a blotting method according to claim 7, wherein in the step S4, data in a mass spectrometry imaging chart is compared with high-resolution mass spectrometry data of a literature and SmetDB database, and the chemical components in the traditional Chinese medicine decoction pieces to be detected are identified.
9. The mass spectrometry imaging method for analyzing chemical components and spatial distribution of dried decoction pieces of traditional Chinese medicine by blotting according to claim 1, wherein step S5 comprises: and comparing the mass spectrum imaging diagram with the original traditional Chinese medicine section photo and the transfer film photo, and analyzing the space distribution rule of the metabolites in the traditional Chinese medicine decoction pieces to be detected.

Description

Mass spectrum imaging method for analyzing chemical components and spatial distribution of dried traditional Chinese medicine decoction pieces by adopting imprinting method Technical Field The invention belongs to the technical field of mass spectrometry imaging, and relates to a mass spectrometry imaging method for analyzing chemical components and spatial distribution of dried traditional Chinese medicine decoction pieces by adopting a blotting method. Background The traditional Chinese medicine decoction pieces are used as traditional Chinese medicine materials capable of being directly blended and prepared, and are key links for ensuring the safety and effectiveness of clinical application of traditional Chinese medicines in the traditional Chinese medicine industry. The research on the chemical component information of the traditional Chinese medicine decoction pieces has important significance for exploring the growth and development of the traditional Chinese medicine and clarifying the pharmacodynamic substance basis and pharmacological action mechanism of the traditional Chinese medicine. Because the metabolites can continuously generate complex interactions in the growth and storage processes of the traditional Chinese medicine, the space-time information of the traditional Chinese medicine needs to be explored, and the spatial distribution of the metabolites in the traditional Chinese medicine decoction piece sample is comprehensively known. Compared with a classical histochemical method, the mass spectrum imaging technology does not need to crush samples, can provide extra space dimension information for metabolite research, and realizes the visual research of traditional Chinese medicine metabolites. Meanwhile, the mass spectrum imaging is a label-free technology, can be used under the condition of not knowing an analyte, and can detect hundreds of compounds at the same time, and compared with the technologies such as a fluorescence microscope, in-situ hybridization and the like, the method has the advantages of high information throughput, high analysis speed and the like. The aerodynamic-assisted desorption electrospray mass spectrometry imaging (AFADESI-MSI) is a high-sensitivity, high-coverage and high-specificity mass spectrometry imaging method, can keep the tissue state intact, has good performance at low m/z value, and provides a powerful tool for the visual analysis of metabolites of Chinese medicinal decoction pieces. The AFADESI-MSI technology greatly promotes the research progress in aspects of traditional Chinese medicine ingredient distribution research, traditional Chinese medicine quality markers, pharmacological and toxicological functions and migration pathways of traditional Chinese medicine metabolites and the like. However, most of traditional Chinese medicine decoction pieces are in a dry state after processing, and are difficult to prepare by adopting a traditional frozen section method, meanwhile, the morphology and physical properties of the traditional Chinese medicine decoction pieces are greatly different, the irregular surface of the traditional Chinese medicine decoction pieces can cause severe change of signal intensity, and a direct imaging method cannot be adopted for preparing samples. The dilemma in the aspect of sample preparation makes the mass spectrum imaging analysis of the traditional Chinese medicine decoction pieces greatly lag behind the tissues of common plants and animals. Disclosure of Invention In order to achieve the above object, the present invention provides the following technical solutions: Step S1, applying pressure to the traditional Chinese medicine decoction pieces to be detected to obtain a relatively flat section; s2, establishing a sandwich structure, and transferring the components of the cross section of the traditional Chinese medicine decoction pieces onto a transfer film by adopting a blotting method; S3, performing ionization scanning on the transfer film in a mass spectrometer by adopting a mass spectrometry imaging detection method to obtain mass spectrometry imaging data of chemical components in the transfer film of the traditional Chinese medicine decoction pieces to be detected; s4, carrying out data processing on the mass spectrum imaging data, and identifying chemical components in the sample to be detected; And S5, analyzing the spatial distribution rule of metabolites in the traditional Chinese medicine decoction pieces to be detected. Preferably, the specific method of step S1 is: Selecting Chinese medicinal decoction pieces with relatively flat cross section and thinner thickness, and applying pressure of 0.5 ton by using a manual tablet pressing machine for 10s to obtain Chinese medicinal decoction pieces with flat cross section to be detected. Preferably, the transfer method of step S2 is specifically: spraying 300-500 mu L of 80% acetonitrile solution on a transfer printing film (nylon microporous filter film), pl