CN110095524B - Triterpene saponin mass spectrum structure analysis method - Google Patents
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Abstract
The invention relates to a mass spectrum structure analysis method of triterpenoid saponin, which comprises the steps of carrying out secondary mass spectrum analysis on a sample under a positive ion mode under different collision energies, then carrying out secondary mass spectrum analysis under a negative ion mode under different collision energies, and carrying out sugar chain composition and structure analysis of a binding site of a triterpenoid saponin compound by utilizing secondary mass spectrum sub-ion fragment characteristics induced by different collision energies under the positive ion and negative ion modes. Can quickly realize accurate identification of the triterpene saponin glycosyl and the binding site.
Description
Technical Field
The invention belongs to the field of separation analytical chemistry, and particularly relates to a mass spectrum structure analysis method of triterpenoid saponin.
Background
The information in this background section is only for enhancement of understanding of the general background of the invention and is not necessarily to be construed as an admission or any form of suggestion that this information forms the prior art that is already known to a person of ordinary skill in the art.
The triterpene saponin compounds are glycoside natural compounds composed of triterpene mother nucleus and glycosyl, and have effects of resisting tumor, resisting aging, relieving fatigue, and regulating immunity. The compounds are widely present in plants of the genus Panax, such as Panax ginseng, Panax quinquefolium, etc. Triterpene saponin has complex sugar chain structure, including hexose, pentose, deoxy hexose, etc., and commonly comprises glucose, galactose, rhamnose, arabinose, xylose, glucuronic acid, galacturonic acid, etc. The common triterpenoid saponin has two sugar chains, and the two sugar chains may be composed of the same glycosyl or different glycosyl and are composed of monosaccharide, disaccharide and trisaccharide. The sugar chains have different binding sites on the triterpene core. The ginseng plants have a large amount of tetracyclic triterpene saponins, common binding sites of the tetracyclic triterpene saponins comprise C-3, C-6 and C-20, and the activity of the tetracyclic triterpene saponins is obviously different due to the difference of the binding sites.
The mass spectrum has high sensitivity and good resolution, and can be widely applied to the analysis of plant triterpenoid saponin compounds. The combination of the ultra-efficient separation capability of liquid chromatography and the mass spectrum technology brings possibility for high-throughput analysis of the triterpenoid saponin. The inventor finds that the triterpene saponin has a complex structure and a plurality of varieties, does not have a commercial standard sample, and brings difficulty to the accurate determination of the sugar chain composition and the binding site. The common secondary mass spectrometry means with single collision energy and single ionization mode cannot accurately identify the combination sites and the sugar chain components, and the multistage mass spectrometry has high requirements on the concentration of compounds, so that the application of the multistage mass spectrometry is limited.
Disclosure of Invention
In view of the problems in the prior art, an object of the present invention is to provide a mass spectrometry structure analysis method for triterpene saponin.
In order to solve the technical problems, the technical scheme of the invention is as follows:
a mass spectrum structure analysis method of triterpenoid saponin comprises the following specific steps: the method comprises the steps of separating a sample by liquid chromatography, carrying out secondary mass spectrometry on the separated sample under positive ion modes under different collision energies, then carrying out secondary mass spectrometry under negative ion modes under different collision energies, and carrying out sugar chain composition and binding site structure analysis on the triterpenoid saponin compound by utilizing secondary mass spectrometry sub-ion fragment characteristics induced by different collision energies under the positive ion and negative ion modes.
The structure and the binding site of glycosyl are accurately determined by mass spectrum characteristics in a positive and negative ion mode and secondary mass spectrum cracking characteristics induced by collision. In the prior art, the combination sites and the sugar chain components cannot be accurately identified by a single collision energy and single ionization mode secondary mass spectrometry, and the low-concentration triterpene saponin cannot be accurately qualitatively analyzed due to higher requirements of multi-stage mass spectrometry on the concentration of compounds. In the invention, the combination of the positive ion mode and the negative ion mode is used for accurately analyzing the structures of the binding sites and the sugar chains respectively. In the present invention, the binding site of the sugar chain is determined by the energy resolution curve of the daughter ion in the positive ion mode. The composition of the sugar group in the sugar chain is determined by the energy resolution curve of the daughter ion in the negative ion mode.
The collision energy range under the positive ion mode is 0-100eV, excluding 0eV, and the interval is 5-10 eV;
the collision energy in the negative ion mode ranges from 0 to 100eV excluding 0eV, and the interval is from 5 to 10 eV.
In some embodiments, the number of collision energy selection points in positive ion mode may be 4-7; in yet other embodiments, 4 collision energies, 40eV, 50eV, 60eV, 70 eV; in yet other embodiments, 5 collision energies, 45eV, 55eV, 65eV, 75eV, and 85eV, respectively; in yet other embodiments, 6 collision energies, 40eV, 50eV, 60eV, 70eV, 80eV, 90eV, respectively; in yet other embodiments, the 7 collision energies are 40eV, 45eV, 50eV, 55eV, 60eV, 65eV, and 70eV, respectively.
In the present invention, binding site information of sugar chains is obtained in a positive ion mode, secondary mass spectrometry is performed at different collision energies, and signals of sugar chains gradually appear as the collision energies increase, and information of sugar chains at different positions is obtained from information of fragment ions. In the positive ion mode of the present invention, as the collision energy increases, the C-O bond at the junction of the sugar chain and the triterpene core is cleaved, and a sodium addition peak of the sugar chain gradually appears, and the sodium addition peak of the sugar chain at the C-20 position appears at a low collision energy and the sodium addition peaks of the sugar chains at the C-3 and C-6 positions appear at a high collision energy.
In some embodiments, 3-5 collision energies in negative ion mode; in yet other embodiments, the 3 collision energies are 65eV, 75eV, and 85eV, respectively; in yet other embodiments, 4 collision energies, 50eV, 60eV, 70eV, and 80eV, respectively; in yet other embodiments, the 5 collision energies are 55eV, 65eV, 75eV, 85eV, and 95eV, respectively.
In the present invention, the sugar chain in the positive ion mode is further analyzed in the negative ion mode to obtain the sugar chain composition. As the energy increases, the sugar group in the sugar chain gradually detaches from the sugar chain, and a specific sugar group molecule can be obtained. Neutral loss of the glycosyl group gradually occurs with the increase of collision energy, and the composition of the glycosyl group within the sugar chain is determined according to the mass number of the neutral loss. Wherein neutral loss 162 is a six carbon sugar, neutral loss 146 is a deoxy six carbon sugar, neutral loss 132 is a five carbon sugar, and neutral loss 176 is a six carbon uronic acid.
In some embodiments, the secondary mass counterion in positive ion mode is [ M + Na]+。
In some embodiments, the secondary mass counterion in the negative ion mode is [ M-H [ ]]-。
In some embodiments, the sample material is panax quinquefolium, panax ginseng, or panax notoginseng.
In some embodiments, the liquid chromatography conditions are: mobile phases a and B were an aqueous solution containing 0.1% formic acid and an acetonitrile solution containing 0.1% formic acid, respectively. The flow rate was 0.3 mL/min. The chromatographic column is a reversed phase C18 chromatographic column. The column temperature is 30-50 ℃. The elution time is 20-50 min. The initial gradient of elution was 5% B and the final gradient was 100% B, and 100% B was used for 3-5 min.
In some embodiments, the sample is obtained by pre-treating the sample material before detection, and the sample is prepared by preparing a solid sample into powder, adding an extracting agent, and performing ultrasonic treatment and centrifugation to obtain a supernatant, namely the sample.
In still other embodiments, the extractant is a mixture of an organic solvent and water, the mass ratio of the organic solvent to the water is 5:1 to 3: 1; in still other embodiments, the organic solvent is one of acetonitrile, methanol, ethanol; in still other embodiments, 1g of powder corresponds to a volume of extractant of 8-12 mL; in still other embodiments, the time of sonication is 20-40 min; in still other embodiments, the time for centrifugation is 7-13 min.
In the invention, a single organic solvent and water are used as an extracting agent, the centrifuged supernatant is used as an extracting solution, and after liquid chromatography, a lot of triterpene saponins with low content and high content are found.
The mass spectrum structure analysis method of the triterpenoid saponin is applied to the characterization of the triterpenoid saponin in the ginseng traditional Chinese medicinal materials.
In some embodiments, the ginseng genus of Chinese medicinal materials includes ginseng, American ginseng, and pseudo-ginseng.
The invention has the beneficial effects that:
according to the invention, the sugar chain composition and the structure analysis of the binding site of the triterpenoid saponin are realized through secondary mass spectrum characteristics under positive ion and negative ion modes with different collision energies;
according to the preparation method of the extracting solution, the organic solvent is one of acetonitrile, methanol and ethanol for extraction, the extracting solution is separated by liquid chromatography, and then secondary mass spectrometry is performed, so that the sensitivity and accuracy of detection and analysis of triterpenoid saponin are improved;
the method is simple and rapid to operate and high in accuracy.
Drawings
The accompanying drawings, which are incorporated in and constitute a part of this specification, are included to provide a further understanding of the invention, and are incorporated in and constitute a part of this specification, illustrate embodiments of the invention and together with the description, serve to explain the invention and not to limit the invention.
FIG. 1 is an energy-resolving curve of a daughter ion of triterpene saponin Re in the positive ion mode in example 1;
FIG. 2 is an energy-resolved curve of a daughter ion of triterpene saponin Re in the anion mode in example 1.
FIG. 3 shows triterpene saponin Rb in positive ion mode in example 22The energy resolution curve of the daughter ion of (a);
FIG. 4 shows triterpene saponin Rb in anion mode in example 22Energy resolution curve of the daughter ion.
FIG. 5 shows triterpene saponin Rb in positive ion mode in example 31The energy resolution curve of the daughter ion of (a);
FIG. 6 shows the triterpene saponin Rb in anion mode in example 31Energy resolution curve of the daughter ion.
Detailed Description
It is to be understood that the following detailed description is exemplary and is intended to provide further explanation of the invention as claimed. Unless defined otherwise, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this invention belongs.
It is noted that the terminology used herein is for the purpose of describing particular embodiments only and is not intended to be limiting of example embodiments according to the present application. As used herein, the singular forms "a", "an" and "the" are intended to include the plural forms as well, and it should be understood that when the terms "comprises" and/or "comprising" are used in this specification, they specify the presence of stated features, steps, operations, devices, components, and/or combinations thereof, unless the context clearly indicates otherwise. The invention will be further illustrated by the following examples
Example 1
Characterization of triterpenoid saponins in American ginseng:
(1) preparing American ginseng extract
Adding 1g radix Panacis Quinquefolii powder into 10mL methanol/water (v/v,4:1) solution, ultrasonic treating for 30min, centrifuging for 10min, and collecting supernatant.
(2) Second order mass spectrometry
Subjecting the supernatant to liquid chromatography-mass spectrometry (UHPLC-Q-TOF-MS) with Acquisty BEH C18(2.1 × 100mm,1.7 μm, Waters) as chromatographic column, mobile phases A and B being 0.1% formic acid in water and 0.1% formic acid in acetonitrile respectively, column temperature 40 deg.C, and elution gradient of 0min and 5% B; 10min, 50% B; 25min, 100% B; 30min, 100% B. The mass spectrum drying gas is 8L/min, the drying gas temperature is 200 ℃, the atomizing gas is 2bar, the capillary voltage is 3500V in a positive ion mode, 3000V in a negative ion mode, the transmission time of a collision cell is 80 mus, RF is 750Vpp, the isolation width is 4, and the mass number range is 50-1200.
Secondary mass spectrometry with different collision energies was performed, with collision energies in positive ion mode of 40eV, 50eV, 60eV, and 70 eV. The collision energy in the negative ion mode was 50eV, 60eV, 70eV, and 80 eV.
(3) Characterization of triterpene saponin:
by daughter ions in positive and negative ion modesThe energy resolution curve qualitatively identifies 35 triterpene saponin components in the American ginseng, and determines glycosyl connecting sites and glycosyl compositions thereof. Taking Re as an example. As shown in FIGS. 1 and 2, Re in the positive ion mode, the signal of the sugar chain 203 daughter ion appears at low energy, and the signal of the sugar chain 349 daughter ion appears at high energy, indicating that the sugar chain 203 is at the C-20 position and the sugar chain 349 is at the C-6 position. In negative ion mode, daughter ion 783([ M-H-162) appears]-)、799([M-H-146]-)、637([M-H-162-146]-)、475([637-162]-). This indicates that the sugar chain 203 is a six-carbon sugar. The sugar chains 349 are one deoxy-six carbon sugar and one six carbon sugar. Wherein M refers to the molecular weight of Re, 783([ M-H-132)]-) A daughter ion obtained by neutral loss of a six-carbon sugar for the sugar chain of Re, 799([ M-H-146 ]]-) Neutral loss of a daughter ion derived from a deoxyhexose for the sugar chain of Re, 637([ M-H-162-]-) Neutral loss of one six-carbon sugar and one deoxy-six-carbon sugar for the sugar chain of Re, 475([ 637-162-)]-) One of the resulting daughter ions of the six carbon sugar is then lost for neutrality.
Example 2
Characterization of triterpene saponins in ginseng:
(1) preparing Ginseng radix extractive solution
Adding 1g Ginseng radix powder into 11mL methanol/water (v/v,4:1) solution, ultrasonic treating for 35min, centrifuging for 12min, and collecting supernatant.
(2) Second order mass spectrometry
Subjecting the supernatant to liquid chromatography-mass spectrometry (UHPLC-Q-TOF-MS) with ZORBAX-SB-AQ C18(2.1 × 100mm,1.8 μm, Agient) as chromatographic column, mobile phases A and B being 0.1% formic acid in water and 0.1% formic acid in acetonitrile respectively, column temperature of 45 deg.C, and elution gradient of 0min and 5% B; 6min, 40% B; 20min, 100% B; 25min, 100% B. The mass spectrum drying gas is 8L/min, the drying gas temperature is 200 ℃, the atomizing gas is 2bar, the capillary voltage is 3500V in a positive ion mode, 3000V in a negative ion mode, the transmission time of a collision cell is 80 mus, RF is 750Vpp, the isolation width is 4, and the mass number range is 50-1200.
Secondary mass spectrometry was performed at different collision energies, 45eV, 55eV, 65eV, 75eV, and 85eV in the positive ion mode. The collision energy in the negative ion mode was 55eV, 65eV, 75eV, 85eV, and 95 eV.
(3) Characterization of triterpene saponin:
through the energy resolution curve of the daughter ions under the positive ion and negative ion modes, 50 triterpene saponin components are qualitatively obtained in the ginseng, and the glycosyl connecting sites and glycosyl composition are determined. With Rb2For example. As shown in fig. 3 and 4, Rb2In the positive ion mode, the signal of sugar chain 355 molecule ion appears at low energy, and the signal of sugar chain 365 molecule ion appears at high energy, indicating that sugar chain 355 is at C-20 position and sugar chain 365 is at C-3 position. In negative ion mode, daughter ions 945([ M-H-132) appear]-)、915([M-H-162]-)、783([M-H-162-132]-)、621([783-162]-)、459([621-162]-). The sugar chain 365 is illustrated as two six carbon sugars. The sugar chain 355 is a five-carbon sugar and a six-carbon sugar. Wherein M means Rb2Molecular weight of 945([ M-H-132)]-) Is Rb2The sugar chain of (1) has a neutral loss of a five-carbon sugar resulting in a daughter ion of 915([ M-H-162 ]]-) Is Rb2783([ M-H-162-]-) Is Rb2The sugar chain of (1) neutral loses a five-carbon sugar and a six-carbon sugar resulting in a daughter ion 621([783-]-) The daughter ion obtained for subsequent neutral loss of a six-carbon sugar, 459([621- & 162 ]]-) One of the resulting daughter ions of the six carbon sugar is then lost for neutrality.
Example 3
Characterization of triterpenoid saponins in panax notoginseng:
(1) preparing Notoginseng radix extractive solution
Adding Notoginseng radix powder 1g into 9mL methanol/water (v/v,4:1) solution, ultrasonic treating for 26min, centrifuging for 8min, and collecting supernatant.
(2) Second order mass spectrometry
Subjecting the supernatant to liquid chromatography-mass spectrometry (UHPLC-Q-TOF-MS) with ZORBAX-SB-AQ C18(2.1 × 100mm,1.8 μm, Agient) as chromatographic column, mobile phases A and B being aqueous solution containing 0.1% formic acid and acetonitrile solution containing 0.1% formic acid, respectively, column temperature of 40 deg.C, elution gradient of 0min, 5% B; 7min, 50% B; 21min, 100% B; 25min, 100% B. The mass spectrum drying gas is 8L/min, the drying gas temperature is 200 ℃, the atomizing gas is 2bar, the capillary voltage is 3500V in a positive ion mode, 3000V in a negative ion mode, the transmission time of a collision cell is 80 mus, RF is 750Vpp, the isolation width is 4, and the mass number range is 50-1200. Secondary mass spectrometry was performed at different collision energies, 40eV, 50eV, 60eV, 70eV, 80eV, and 90eV in the positive ion mode. The collision energy in the negative ion mode was 65eV, 75eV, and 85 eV.
(3) Characterization of triterpene saponin:
through the energy resolution curve of the daughter ions under the positive ion and negative ion modes, 20 triterpene saponin components are qualitatively obtained in the pseudo-ginseng, and the glycosyl connecting sites and glycosyl composition are determined. With Rb1For example. As shown in fig. 5 and 6, Rb1In the positive ion mode, only the signal of the sugar chain 365 molecule ion appears, and the relative intensity is always 100% when the collision energy is 60eV or more. In negative ion mode, daughter ion 945([ M-H-162)]-)、783([M-H-162-162]-)、621([783-162]-)、459([621-162]-). The sugar chain 365 is illustrated as two six carbon sugars. And the C-20 position and the C-3 position are sugar chains of two six-carbon sugars. Wherein M means Rb1Molecular weight of 945([ M-H-162)]-) Is Rb1783([ M-H-162-]-) Is Rb1The sugar chain of (1) has a neutral loss of two six-carbon sugar derived daughter ions 621([783-]-) The daughter ion obtained for subsequent neutral loss of a six-carbon sugar, 459([621- & 162 ]]-) One of the resulting daughter ions of the six carbon sugar is then lost for neutrality.
Example 4
Characterization of triterpenoid saponins in American ginseng:
(1) preparing American ginseng extract
Adding 1g radix Panacis Quinquefolii powder into 8mL methanol/water (v/v,4:1) solution, ultrasonic treating for 25min, centrifuging for 13min, and collecting supernatant.
(2) Second order mass spectrometry
Subjecting the supernatant to liquid chromatography-mass spectrometry (UHPLC-Q-TOF-MS) with Acquisty BEH C18(2.1 × 100mm,1.7 μm, Waters) as chromatographic column, mobile phases A and B being 0.1% formic acid in water and 0.1% formic acid in acetonitrile respectively, column temperature 40 deg.C, and elution gradient of 0min and 5% B; 10min, 50% B; 25min, 100% B; 30min, 100% B. The mass spectrum drying gas is 8L/min, the drying gas temperature is 200 ℃, the atomizing gas is 2bar, the capillary voltage is 3500V in a positive ion mode, 3000V in a negative ion mode, the transmission time of a collision cell is 80 mus, RF is 750Vpp, the isolation width is 4, and the mass number range is 50-1200. Secondary mass spectrometry was performed at different collision energies, 40eV, 45eV, 50eV, 55eV, 60eV, 65eV, and 70eV in the positive ion mode. The collision energy in the negative ion mode was 50eV, 60eV, 70eV, and 80 eV.
(3) Characterization of triterpene saponin:
through the energy resolution curves of the daughter ions under the positive ion and negative ion modes, 35 triterpene saponin components are qualitatively obtained in the American ginseng, and glycosyl connecting sites and glycosyl compositions of the triterpene saponin components are determined. With Rb2For example. Rb2In the positive ion mode, the signal of the sugar chain 355 appears at a low energy, and the signal of the sugar chain 365 appears at a high energy, indicating that the sugar chain 355 is at the C-20 position and the sugar chain 365 is at the C-3 position. In negative ion mode, daughter ions 945([ M-H-132) appear]-)、915([M-H-162]-)、783([M-H-162-132]-)、621([783-162]-)、459([621-162]-). The sugar chain 365 is illustrated as two six carbon sugars. The sugar chain 355 is a five-carbon sugar and a six-carbon sugar. Wherein M means Rb2Molecular weight of (2).
Comparative example 1
Characterization of triterpenoid saponins in American ginseng:
(1) preparing American ginseng extract
Adding 1g radix Panacis Quinquefolii powder into 10mL methanol/water (v/v,4:1) solution, ultrasonic treating for 30min, centrifuging for 10min, and collecting supernatant.
(2) Second order mass spectrometry
Subjecting the supernatant to liquid chromatography-mass spectrometry (UHPLC-Q-TOF-MS) with Acquisty BEH C18(2.1 × 100mm,1.7 μm, Waters) as chromatographic column, mobile phases A and B being 0.1% formic acid in water and 0.1% formic acid in acetonitrile respectively, column temperature 40 deg.C, and elution gradient of 0min and 5% B; 10min, 50% B; 25min, 100% B; 30min, 100% B. The mass spectrum drying gas is 8L/min, the drying gas temperature is 200 ℃, the atomizing gas is 2bar, the capillary voltage is 3500V in a positive ion mode, 3000V in a negative ion mode, the transmission time of a collision cell is 80 mus, RF is 750Vpp, the isolation width is 4, and the mass number range is 50-1200. Secondary mass spectrometry with different collision energies was performed, and the collision energies in the positive ion mode were 70eV, 80eV, 90eV, and 100 eV. The collision energy in the negative ion mode was 40eV, 50eV, and 60 eV.
(3) Characterization of triterpene saponin:
through the energy resolution curve of the daughter ions under the positive ion and negative ion modes, only 15 triterpene saponin components in the American ginseng determine the glycosyl connecting sites and glycosyl composition. Because the collision energy is not properly selected under the positive ion and negative ion modes, the glycosyl connecting sites and the sugar chain components cannot be determined by 20 triterpene saponin components. The collision energy in the positive ion mode is too high, so that the discrimination of sugar chain molecule ions becomes poor. For example, Re, in which the relative strengths of the sugar chains 349 and 203 are high at collision energies of 70eV to 100eV, the problem of cleavage cannot be effectively distinguished, so that the position of linkage of the sugar chain to the aglycone cannot be determined. The collision energy in the negative ion mode is too low to obtain rich data of the fragment of the daughter ion. Such as Rb2Wherein the daughter ion 621 which has a neutral loss of three saccharides and the daughter ion 459 which has a neutral loss of four saccharides are not obtained at a collision energy of 40-60eV, the composition of the sugar chain thereof cannot be determined efficiently.
Comparative example 2
Characterization of triterpenoid saponins in American ginseng:
(1) preparing American ginseng extract
Adding 1g radix Panacis Quinquefolii powder into 10mL methanol/water (v/v,4:1) solution, ultrasonic treating for 30min, centrifuging for 10min, and collecting supernatant.
(2) Second order mass spectrometry
Subjecting the supernatant to liquid chromatography-mass spectrometry (UHPLC-Q-TOF-MS) with Acquisty BEH C18(2.1 × 100mm,1.7 μm, Waters) as chromatographic column, mobile phases A and B being 0.1% formic acid in water and 0.1% formic acid in acetonitrile respectively, column temperature 40 deg.C, and elution gradient of 0min and 5% B; 10min, 50% B; 25min, 100% B; 30min, 100% B. The mass spectrum drying gas is 8L/min, the drying gas temperature is 200 ℃, the atomizing gas is 2bar, the capillary voltage is 3500V in a positive ion mode, 3000V in a negative ion mode, the transmission time of a collision cell is 80 mus, RF is 750Vpp, the isolation width is 4, and the mass number range is 50-1200. Secondary mass spectrometry was performed in positive ion mode at different collision energies, 50eV, 60eV, 70eV, 80eV, and 90 eV.
(3) Characterization of triterpene saponin:
through the energy resolution curve of the daughter ions under the positive ion mode, only the connection sites of the sugar chains and aglycones of the triterpenoid saponin components can be identified in the structural analysis process of the American ginseng triterpenoid saponin, and the specific glycosyl compositions in the sugar chains cannot be identified.
The above description is only a preferred embodiment of the present invention and is not intended to limit the present invention, and various modifications and changes may be made by those skilled in the art. Any modification, equivalent replacement, or improvement made within the spirit and principle of the present invention should be included in the protection scope of the present invention.
Claims (21)
1. A mass spectrum structure analysis method of triterpenoid saponin is characterized by comprising the following steps: the method comprises the following specific steps: separating a sample by liquid chromatography, performing secondary mass spectrometry on the separated sample under positive ion modes under different collision energies, performing secondary mass spectrometry under negative ion modes under different collision energies, and performing sugar chain composition and binding site structure analysis on the triterpenoid saponin compound by using secondary ion characteristics induced by different collision energies under the positive ion and negative ion modes;
the collision energy range under the positive ion mode is 0-100eV, excluding 0eV, and the interval is 5-10 eV;
the collision energy range under the negative ion mode is 0-100eV, excluding 0eV, and the interval is 5-10 eV; the number of collision energy selection points in the positive ion mode is 4-7; determining the binding sites of the sugar chains through an energy resolution curve of the daughter ions in a positive ion mode; the composition of the sugar group in the sugar chain is determined by the energy resolution curve of the daughter ion in the negative ion mode.
2. The method of claim 1, wherein: in positive ion mode, 4 collision energies, 40eV, 50eV, 60eV, and 70 eV.
3. The method of claim 1, wherein: in positive ion mode, 5 collision energies, 45eV, 55eV, 65eV, 75eV, and 85 eV.
4. The method of claim 1, wherein: in the positive ion mode, 6 collision energies were 40eV, 50eV, 60eV, 70eV, 80eV, and 90eV, respectively.
5. The method of claim 1, wherein: in the positive ion mode, 7 collision energies were 40eV, 45eV, 50eV, 55eV, 60eV, 65eV, and 70eV, respectively.
6. The method of claim 1, wherein: 3-5 collision energies in negative ion mode.
7. The method of claim 6, wherein: the collision energies of 3 were 65eV, 75eV, and 85eV, respectively.
8. The method of claim 6, wherein: the 4 collision energies were 50eV, 60eV, 70eV, and 80eV, respectively.
9. The method of claim 6, wherein: 5 collision energies, 55eV, 65eV, 75eV, 85eV, and 95 eV.
10. The method of claim 1, wherein: the secondary mass spectrum ion under the positive ion mode is [ M + Na]+。
11. The method of claim 1, wherein: in negative ion modeThe secondary mass spectrum ion is [ M-H ]]-。
12. The method of claim 1, wherein: the sample material is radix Panacis Quinquefolii, Ginseng radix or Notoginseng radix.
13. The method of claim 1, wherein: liquid chromatography conditions: mobile phases a and B are an aqueous solution containing 0.1% formic acid and an acetonitrile solution containing 0.1% formic acid, respectively; the flow rate is 0.3 mL/min; the chromatographic column is a reversed phase C18 chromatographic column; the column temperature is 30-50 ℃; the elution time is 20-50 min; the initial gradient of elution was 5% B and the final gradient was 100% B, and 100% B was used for 3-5 min.
14. The method of claim 1, wherein: the sample is obtained by preprocessing the sample raw material before detection, and the preparation method of the sample comprises the steps of preparing a solid sample into powder, adding an extracting agent for dissolution, and carrying out ultrasonic treatment and centrifugation to obtain supernatant, namely the sample.
15. The method of claim 14, wherein: the ultrasonic treatment time is 20-40 min.
16. The method of claim 14, wherein: the centrifugation time is 7-13 min.
17. The method of claim 14, wherein: 1g of powder corresponds to a volume of 8-12mL of extractant.
18. The method of claim 14, wherein: the extractant is a mixture of an organic solvent and water, and the mass ratio of the organic solvent to the water is 5:1-3: 1.
19. The method of claim 18, wherein: the organic solvent is one of acetonitrile, methanol and ethanol.
20. Use of the method of any one of claims 1-19 for structural resolution of triterpenoid saponins in a ginseng plant.
21. Use according to claim 20, characterized in that: the Panax Chinese medicinal materials include Ginseng radix, radix Panacis Quinquefolii and Notoginseng radix.
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