CN115166068B - Temperature-internal medicine sharing and differential volatile component analysis based on headspace-gas chromatography-ion mobility spectrometry - Google Patents
Temperature-internal medicine sharing and differential volatile component analysis based on headspace-gas chromatography-ion mobility spectrometry Download PDFInfo
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Abstract
The invention belongs to the field of traditional Chinese medicine quality evaluation, and particularly relates to temperature lining medicine sharing and differential volatile component analysis based on headspace-gas chromatography-ion mobility spectrometry. The method takes black pepper, fennel, pricklyash peel, dried ginger, cinnamon and clove with relatively high volatile oil content in the interior-warming medicine as a research object, adopts a gas phase ion mobility spectrometry (GC-IMS) technology to carry out statistics and analysis on volatile components, has rapid and sensitive detection process, can provide certain data support for analyzing the pharmacodynamic substance basis of the interior-warming traditional Chinese medicine by carrying out different analysis on the organic volatile components of each traditional Chinese medicine, and simultaneously provides certain reference for quality evaluation of the traditional Chinese medicine serving as food application because the traditional Chinese medicine is a flavoring spice with relatively high daily diet use frequency.
Description
Technical Field
The invention belongs to the field of traditional Chinese medicine quality evaluation, and particularly relates to temperature lining medicine sharing and differential volatile component analysis based on headspace-gas chromatography-ion mobility spectrometry.
Background
The interior-warming herbs are common clinical herbs and have a long history of administration. The principle of the traditional Chinese medicine is that the cold is the heat of the cold type and the cold of the heat type is the basic principle of the clinical science, so that the medicine is mainly used for treating the interior cold syndrome and is widely applied to the diseases of the digestive system and the cardiovascular system clinically. In recent years, with the continuous and deep research on chemical components and pharmacology of the interior-warming drugs, the effects of resisting tumor, resisting bacteria, resisting inflammation, easing pain and the like are also increasingly focused by people, and meanwhile, a plurality of interior-warming traditional Chinese medicines are important sources of kitchen seasonings, and play roles in regulating body, preventing diseases and building up body in daily diet.
The interior-warming herbs have pungent taste, and modern researches suggest that the pungent taste is related to the volatile ingredients contained in the herbs, which can manifest their pungent and dispersed properties, so the volatile ingredients are also considered as the main active ingredients of the interior-warming herbs. Modern researches have shown that the interior-warming herbs contain volatile oil components except aconite root, and the content is not equal but less than 1% to 20%. As a Chinese medicine with similar efficacy, the interior-warming herbs have the same pharmacodynamic substance basis, and the difference components determine the difference between the pharmacodynamic actions. However, at present, no statistical analysis is known on the composition of volatile components among interior-warming traditional Chinese medicines. Meanwhile, in the research of the volatile components of single medicine at present, the steam distillation method is mostly adopted for extraction, but the method has low volatile oil yield, long extraction process time and difficult popularization, and meanwhile, the density and the yield of different traditional Chinese medicine volatile oils have great differences, so that the parallel operation of the extraction process is difficult to realize.
The headspace ion gas chromatography is a novel technology based on gas chromatography and ion migration chromatography, and the sample is not required to be pretreated when the fingerprint characteristics of the volatile compounds are detected, so that the original appearance of the sample can be maintained. The basic principle of ion migration is that ions have different migration speeds when passing through an electric field under normal pressure to realize separation.
Disclosure of Invention
According to the invention, black pepper, fennel, pricklyash peel, dried ginger, cinnamon and clove with relatively high volatile oil content in the interior-warming medicine are taken as research objects, the volatile components are counted and analyzed by adopting a gas phase ion mobility spectrometry (GC-IMS) technology, the detection process is rapid and sensitive, a certain data support can be provided for analyzing the drug effect substance basis of the interior-warming medicine through the different analysis of the organic volatile components of each traditional Chinese medicine, and meanwhile, the traditional Chinese medicine is a flavoring spice with relatively high daily diet use frequency, so that a research result can also provide a certain reference for the quality evaluation of the traditional Chinese medicine as food application.
The technical scheme of the invention is as follows:
temperature lining medicine sharing and differential volatile component analysis based on headspace-gas chromatography-ion mobility spectrometry comprises the following steps:
(1) Taking coarse powder of the interior medicine in temperature, placing the coarse powder in a headspace sampling bottle, incubating, testing by using a gas phase ion mobility spectrometer, and parallelly measuring each sample;
(2) Qualitative analysis is carried out on the substances by using an NIST database and an IMS database; drawing gas-phase ion mobility spectrometry fingerprint patterns of all decoction pieces by adopting a Gallery Plot plug-in of software Laboratory Analytical Viewer, and visually and quantitatively comparing the differences of volatile organic compounds among all samples; dynamic principal component analysis (Dynamic PCA) software is used for carrying out cluster analysis and similarity analysis on the sample and determining the types of unknown components; the euclidean distance matrix between samples is calculated using "nearest neighbor" fingerprint analysis, the "nearest neighbor" is found by retrieving the minimum distance, and relatively tight population measurements are observed compared to more distant populations.
Preferably, the setting conditions of the gas phase ion mobility spectrometer in the step (1) are: MXT-5 column 15 m,0.53 mm; carrier gas/drift gas: nitrogen gas; carrier gas flow rate: carrier gas flow rate: 0-2 min,2 mL -1 ；2～20 min，2～100 mL•min -1 ，20～50 min，100 mL•min -1 The method comprises the steps of carrying out a first treatment on the surface of the Drift airflow volume: 150 mL.min -1 The method comprises the steps of carrying out a first treatment on the surface of the Column temperature: 60. the temperature is lower than the temperature; IMS temperature: 45. the temperature is lower than the temperature; sample injection needle temperature: 85. the temperature is lower than the temperature; sample injection volume: 100. mu L.
Further, the common volatile components are: limonene, alpha-phellandrene, pseudolimonene, alpha-myrcene, alpha-pinene, delta-2-carene, sabinene, acetone, ethanol, (E) -beta-ocimene.
Further, the relative contents of 10 common components in each traditional Chinese medicine are respectively as follows: 31.05% of black pepper, 38.25% of cinnamon, 20.11% of pepper, 18.42% of clove, 23.64% of dried ginger and 21.00% of fennel.
The beneficial effects of the invention are that
The invention establishes the GC-IMS characteristic spectrum of six interior-warming traditional Chinese medicines, and totally defines 80 volatile components, wherein 10 common components with similar content are respectively provided with 6-24 different characteristic volatile components. The 10 common components provide a certain data support for screening the effective components of the interior warming medicine, so that the composition can be used as target components for further activity evaluation research. Meanwhile, the unique volatile components of different interior-warming herbs can be used for distinguishing interior-warming herbs. The clustering analysis result shows that the samples between the black pepper, the dried ginger and the cinnamon which are both hot have higher similarity and can be obviously distinguished from the fennel, the pricklyash peel and the clove which are both warm, so the method provides a certain thought for the characteristic analysis of the drug property of the volatile traditional Chinese medicine. In addition, the headspace-gas chromatography-ion mobility spectrometry technology is combined with the principal component analysis method to realize the rapid identification of volatile components in the Chinese medicine in temperature, and has good application prospect in the analysis of volatile components.
Drawings
FIG. 1 is a sample gas phase ion mobility spectrometry;
FIG. 2 is a graph showing the difference in ion mobility chromatography for each piece subtracted with black pepper as a reference;
FIG. 3 is a Gallery Plot of volatile organic compounds selected from a gas phase ion mobility spectrometry; and (3) injection: each row of the map represents one sample (HHJ No. 1-3, HJ No. 4-6, RG No. 7-9, DX No. 10-12, GJ No. 13-15, XHX No. 16-18 in order from top to bottom, each sample was assayed in triplicate). Each column represents a signal peak (numbers 1 to 80 correspond to the numbers in table 1, and 1 'to 40' are unknown components). Region a is 42, 43, 16, 15, 17, 18, 21, 19, 22, 68, 1' in sequence from left to right; b is 14, 2', 3', 39, 52, 4' in sequence; c is 29, 30, 5', 6, 60, 67, 69 in sequence; d is 23, 20, 46, 47, 24, 25, 7', 4, 8', 7, 12, 5, 13, 9', 10', 65, 64, 50, 11', 12', 13', 14', 15', 70, 16', 17', 18', 73, 44, 72, 28, 74, 19', 20', 75, 21', 76, 78, 22', 23', 24', 77 in that order; e is 1, 25', 26', 8, 26, 27, 45, 37, 38, 48, 49, 53, 62, 59, 27', 28', 29', 30', 31' in order; f is 32', 2, 3, 32, 51, 33, 34, 35, 36, 40, 41, 33', 34', 66, 35', 71, 79, 80 in that order; g is 61, 11, 36', 9, 10, 37', 56, 57, 31, 54, 55, 63, 58, 38', 39', 40' in that order;
FIG. 4 is a PCA analysis of all samples;
fig. 5 is a graph of euclidean distance between samples.
Detailed Description
Example 1
Taking 1g of each of the coarse powder samples, placing the coarse powder samples into a 20mL headspace sample injection bottle, incubating at 60 ℃ for 15 minutes, and testing by using a gas phase ion mobility spectrometer FlavourSpec for headspace sample injection, wherein each sample is measured three times in parallel.
MXT-5 column (15 m,0.53 mm). Carrier gas/drift gas: nitrogen gas. Carrier gas flow rate: carrier gas flow rate: 0-2 min,2 mL.min-1;2 to 20 min,2 to 100 mL -1 ，20～50 min，100 mL•min -1 . Drift airflow volume: 150 mL.min -1 . Column temperature: 60. DEG C. IMS temperature: 45. DEG C. Sample injection needle temperature: 85. DEG C. Sample injection volume: 100. mu L.
2.3 Data analysis
Qualitative analysis of the material was performed using a NIST database and an IMS database built in the software. And drawing GC-IMS fingerprint patterns of all decoction pieces by adopting a gally Plot plug-in of software Laboratory Analytical Viewer (LAV), and visually and quantitatively comparing the difference of volatile organic compounds among all samples. Dynamic PCA (Dynamic principal component analysis) software is used to perform cluster analysis and similarity analysis on samples, and to determine unknown component types. The euclidean distance matrix between samples is calculated using "nearest neighbor" fingerprint analysis, the "nearest neighbor" is found by retrieving the minimum distance, and relatively tight population measurements are observed compared to more distant populations.
3. Results and discussion
3.1 Gas phase ion mobility spectrometry analysis sample gas phase ion mobility spectrometry is shown in figure 1, where each point on either side of the RIP (red vertical line at abscissa 1.0, drift time about 7.97 ms) represents a volatile organic compound. Color represents the concentration of the substance, white represents less concentration, red represents greater concentration, and darker color represents greater concentration. The black pepper (HHJ) spectrum is taken as a reference, the other sample spectrum is deducted and reduced to be a reference, the consistent part of the black pepper (HHJ) spectrum and the reference is deducted and reduced to be white, red represents that the concentration of the substance is higher than that of the reference, and blue represents that the concentration of the substance is lower than that of the reference, and the result is shown in figure 2. As can be seen, there is some similarity in composition between the different samples, as well as a more distinct difference.
GC-IMS characteristic spectrum of each Chinese medicinal decoction piece
The HS-GC-IMS characteristic patterns of the decoction pieces of black pepper, pricklyash peel and the like are shown in figure 3 (the labeled peak numbers in the figure are consistent with Table 1), and the figure 3 contains all signal peaks which can be detected under the detection condition, so that the composition and the relative content of volatile substances of different samples can be intuitively represented. As can be seen from the figure, the Chinese medicines in each temperature category contain common Volatile Organic Compounds (VOCs), have certain similarity, and are shown in yellow boxes to contain common and similar components in different samples. The positions, the numbers, the intensities and the time of the ion peaks of different decoction pieces also show certain differences, and substances shown by red boxes in the figure are volatile components of characteristics in each sample, and the relative content of the substances in each sample is obviously higher than that of other samples, especially the Chinese prickly ash (D area in the figure) contains a large amount of characteristic volatile components.
Table 1 qualitative analysis of volatile compounds
The results of qualitative analysis of volatile compounds in the samples are shown in Table 1. The total of 80 components are identified, wherein, the components are terpenes 16, alcohols 14, ketones 12, aldehydes 23, acids 1, ethers 3, esters 9 and aromatic hydrocarbons 2. Due to the presence of monomers, dimers and even polymers, several single compounds may produce multiple signals or spots due to their different concentrations. Monomers and dimers are represented by M and D, respectively, in table 1.
The total of 6 Chinese medicinal materials contains 10 similar components, wherein the 10 known components comprise 8 terpene components, 1 ketone component, and alcohols1 component. The relative contents of the 10 common components in each traditional Chinese medicine are respectively as follows: 31.05% of black pepper, 38.25% of cinnamon, 20.11% of pepper, 18.42% of clove, 23.64% of dried ginger and 21.00% of fennel. In terms of content, the common components are of significance for efficacy analysis. Among the 10 common components, limonene is a natural functional monoterpene and has good antibacterial, anti-inflammatory, cough and asthma relieving effects. Myrcene and laureneα-pinene has good anti-inflammatory, antioxidant and cytoprotective effects, and ocimene can inhibit candida albicans activity, and the components may be common medicinal components of anti-inflammatory and anti-diarrhea of the interior warming medicine.
Claims (1)
1. The temperature lining medicine sharing and differential volatile component analysis based on headspace-gas chromatography-ion mobility spectrometry is characterized by comprising the following steps of:
(1) Taking coarse powder of the interior medicine in temperature, placing the coarse powder in a headspace sampling bottle, incubating, testing by using a gas phase ion mobility spectrometer, and parallelly measuring each sample;
(2) Qualitative analysis is carried out on the substances by using an NIST database and an IMS database; drawing gas-phase ion mobility spectrometry fingerprint patterns of all decoction pieces by adopting a Gallery Plot plug-in of software Laboratory Analytical Viewer, and visually and quantitatively comparing the differences of volatile organic compounds among all samples; dynamic principal component analysis (Dynamic PCA) software is used for carrying out cluster analysis and similarity analysis on the sample and determining the types of unknown components; calculating Euclidean distance matrix between samples by adopting 'nearest neighbor' fingerprint analysis, finding 'nearest neighbor' by searching minimum distance, and observing a relatively compact group measurement result compared with a more distant group;
the setting conditions of the gas phase ion mobility spectrometer in the step (1) are as follows: MXT-5 column 15 m,0.53 mm; carrier gas/drift gas: nitrogen gas; carrier gas flow rate: carrier gas flow rate: 0-2 min,2 mL -1 ；2～20 min，2～100 mL•min -1 ，20～50 min，100 mL•min -1 The method comprises the steps of carrying out a first treatment on the surface of the Drift airflow volume: 150 mL.min -1 The method comprises the steps of carrying out a first treatment on the surface of the Column temperature: 60. the temperature is lower than the temperature; IMS temperature: 45. the temperature is lower than the temperature; sample injection needle temperature: 85. the temperature is lower than the temperature; sample injection volume: 100. mu L;
the common volatile components are: limonene, alpha-phellandrene, pseudolimonene, alpha-myrcene, alpha-pinene, delta-2-carene, sabinene, acetone, ethanol, (E) -beta-ocimene;
taking a black pepper spectrogram as a reference, and deducting the reference from other sample spectrograms;
the relative contents of the 10 common components in each traditional Chinese medicine are respectively as follows: 31.05% of black pepper, 38.25% of cinnamon, 20.11% of pepper, 18.42% of clove, 23.64% of dried ginger and 21.00% of fennel.
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