CN115184503A - Analysis method of tobacco hydroxy geranyl linalool diterpene enol glycoside and two-dimensional liquid chromatography-mass spectrometry combined system - Google Patents
Analysis method of tobacco hydroxy geranyl linalool diterpene enol glycoside and two-dimensional liquid chromatography-mass spectrometry combined system Download PDFInfo
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- OSWFIVFLDKOXQC-UHFFFAOYSA-N 4-(3-methoxyphenyl)aniline Chemical compound COC1=CC=CC(C=2C=CC(N)=CC=2)=C1 OSWFIVFLDKOXQC-UHFFFAOYSA-N 0.000 claims description 13
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- G01N30/00—Investigating or analysing materials by separation into components using adsorption, absorption or similar phenomena or using ion-exchange, e.g. chromatography or field flow fractionation
- G01N30/02—Column chromatography
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- G01N30/00—Investigating or analysing materials by separation into components using adsorption, absorption or similar phenomena or using ion-exchange, e.g. chromatography or field flow fractionation
- G01N30/02—Column chromatography
- G01N30/26—Conditioning of the fluid carrier; Flow patterns
- G01N30/38—Flow patterns
- G01N30/46—Flow patterns using more than one column
- G01N30/461—Flow patterns using more than one column with serial coupling of separation columns
- G01N30/463—Flow patterns using more than one column with serial coupling of separation columns for multidimensional chromatography
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- G01N30/72—Mass spectrometers
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Abstract
The invention provides an analysis method of tobacco hydroxy geranyl linalool diterpene enol glycoside and a two-dimensional liquid chromatography-mass spectrometry combined system. The method comprises the following steps: pretreating tobacco leaves, and qualitatively and quantitatively analyzing samples to be detected obtained by pretreatment by adopting a two-dimensional liquid chromatography-mass spectrometry combined method: separating out a target component through a first-dimension liquid chromatogram, trapping the target component after dilution by a solution supplement, eluting the trapped component, separating through a second-dimension liquid chromatogram, and performing qualitative and quantitative analysis through a mass spectrum. The system comprises a first-dimension liquid chromatogram, a compensation pump, a communication interface, a switching valve, a trapping column, a second-dimension liquid chromatogram, a mass spectrum and an ultraviolet detector. The invention has high sensitivity and good accuracy, and realizes the semi-quantitative determination of 7 HGL-DTGs.
Description
Technical Field
The invention belongs to the technical field of tobacco chemistry, and relates to an analysis method of tobacco hydroxy geranyl linalool diterpene enol glycoside and a two-dimensional liquid chromatography-mass spectrometry system.
Background
Hydroxygeranyl linalool diterpene enol glycosides (HGL-DTGs) are rich secondary metabolites in tobacco stem and leaf tissues, are mediated through a secondary metabolic pathway in the growth process, and have strong insect resistance. The HGL-DTGs have the following structural formula, and are composed of 1 17-hydroxy geraniol main chain, and 2 different branched groups (R) are respectively formed at the C3 and C17 positions of the main chain with units such as glucose, rhamnose and the like 1 、R 2 ) And R is 2 And the modified HGL-DTGs are modified together at C6 by matching with malonimide, and finally, a plurality of different HGL-DTGs isomers are formed.
As the HGL-DTGs are a group of more complex secondary metabolites of the tobacco, the composition of the tobacco is complex, the matrix effect is obvious, and the accurate determination difficulty of the HGL-DTGs is higher. At present, only a qualitative analysis method of HGL-DTGs is reported, pretreatment steps are complicated, steps such as multi-step enrichment, separation, preparation and the like are involved, and a quantitative and semi-quantitative analysis method of the HGL-DTGs in tobaccos is not reported.
Disclosure of Invention
In view of the defects of the prior art, the invention aims to provide an analysis method of tobacco hydroxy geranyl linalool diterpene enol glycoside and a two-dimensional liquid chromatography-mass spectrometry combined system, which have high sensitivity and good accuracy and realize semi-quantitative determination of 7 HGL-DTGs.
In order to achieve the above objects and other related objects, a first aspect of the present invention provides a method for analyzing tobacco hydroxy-geranyl linalool diterpene enol glycoside, comprising the steps of: pretreating tobacco leaves, and qualitatively and quantitatively analyzing a to-be-detected sample obtained by pretreatment by adopting a two-dimensional liquid chromatography-mass spectrometry combined method: separating out target components through a first-dimension liquid chromatogram, trapping the target components after solution supplementing dilution, eluting trapped components, separating through a second-dimension liquid chromatogram, and performing qualitative and quantitative analysis through a mass spectrum.
Preferably, the pretreatment comprises the following steps:
1) Crushing and grinding the tobacco leaves under liquid nitrogen;
2) Oscillating the tobacco leaves obtained in the step 1) in an organic solvent, centrifuging and then coating a film to obtain a sample to be detected.
Preferably, at least one of the following technical features is also included:
a) The conditions of the first dimension liquid chromatogram are as follows: first dimension liquid chromatography: reversed phase liquid chromatography; a first dimension chromatographic column: CAPCELL PAK MGIII C18 (50 mm. Times.2.0 mm,3 μm); flow rate: 0.05-1 mL/min; column temperature: 20 to 60 ℃; sample introduction amount: 1-20.0 mu L; first dimension mobile phase a phase: water plus 0.05 to 2 percent of formic acid; first dimension mobile phase B phase: methanol and 0.05 to 2 percent of formic acid; gradient leaching;
b) The conditions of the second dimension liquid chromatogram are as follows: second dimension liquid chromatography: reversed phase liquid chromatography; a second dimension chromatographic column: agilent ZORBAX 300SB C18 (2.1 mm. Times.100mm, 3.5 μm); flow rate: 0.05-1 mL/min; column temperature: 20 to 60 ℃; second-dimensional mobile phase a phase: water and 0.05 to 2 percent of formic acid; second-dimensional mobile phase B phase: acetonitrile and 0.05-2% formic acid; and (4) gradient elution.
More preferably, in feature a), the specific procedure of the gradient elution is:
0-5 min, first dimension mobile phase A phase: the volume ratio of the first-dimension mobile phase B phase is 70:30, of a nitrogen-containing gas;
5-20 min, first dimension mobile phase A phase: the volume ratio of the first-dimension mobile phase B phase is 70:30 to 5:95;
20-25 min, first dimension mobile phase A phase: the volume ratio of the first-dimension mobile phase B phase is 5:95;
25-26 min, first dimension mobile phase A phase: the volume ratio of the first-dimensional mobile phase B phase is 5: 95-70: 30, of a nitrogen-containing gas;
26-50 min, first dimension mobile phase A phase: the volume ratio of the first-dimension mobile phase B phase is 70:30.
more preferably, in feature b), the specific procedure of the gradient elution is:
0-22 min, second dimension mobile phase A phase: the volume ratio of the second dimension mobile phase B phase is 95:5;
22-27 min, second dimension mobile phase A phase: the volume ratio of the second-dimensional mobile phase B phase is 95:5 to 70:30;
27-37min, second dimension mobile phase A phase: the volume ratio of the second dimension mobile phase B phase is 70:30 to 40:60, adding a solvent to the mixture;
37-37.1 min, second dimension mobile phase A phase: the volume ratio of the second-dimension mobile phase B phase is 40: 60-5: 95;
37.1-45 min, second dimension mobile phase A phase: the volume ratio of the second-dimension mobile phase B phase is 5:95;
45-45.1 min, second dimension mobile phase A phase: the volume ratio of the second-dimension mobile phase B phase is 5: 95-95: 5;
45.1-50 min, second dimension mobile phase A phase: the volume ratio of the second-dimensional mobile phase B phase is 95:5.
preferably, the trapping conditions are:
a trapping column: thermo scientific Acclaim Polar Advantage II (4.6 mm. Times.10mm, 5 μm);
liquid supplementing: water;
column temperature: 20 to 60 ℃;
cutting time: 16.8min-21.8min;
the flow rate of the liquid supplement is 0-15min, and the flow rate is 0.1-1 mL/min;16-22min, 0.1-1.0 mL/min;23 to 30min,0.1 to 1mL/min.
Preferably, the conditions of the mass spectrum are: electrospray ionization (ESI), negative ion scanning, multi-reaction detection (MRM) mode; electrospray voltage: -3000-6000V; ion source temperature: 200 to 600 ℃; residence monitoring time of ion pair: 10-100 ms; atomizing gas pressure: 20 to 60Psi; auxiliary gas pressure: 20 to 60Psi; mass spectrometry scan time: 27-37min.
Preferably, the tobacco hydroxy geranyl linalool diterpenoid enol glycosides include lycoumoside I, nicotianoside III, nicotianoside IV, nicotianoside V, nicotianoside VI and Nicotianoside VII.
The invention provides a two-dimensional liquid chromatography-mass spectrometry combined system, which comprises a first-dimensional liquid chromatography, a compensation pump, a communication interface, a switching valve, a trapping column, a second-dimensional liquid chromatography, a mass spectrum and an ultraviolet detector, wherein the first-dimensional liquid chromatography is connected with the first-dimensional liquid chromatography;
the first dimension liquid chromatogram comprises a first dimension pump and a first dimension chromatographic column which are communicated, and the second dimension liquid chromatogram comprises a second dimension pump and a second dimension chromatographic column which are communicated;
the communication interface is respectively communicated with the sample outlet end of the first-dimension chromatographic column, the compensation pump and the switching valve through pipelines;
the sample outlet end of the second-dimension chromatographic column is communicated with the mass spectrum;
the switching valve is respectively communicated with the second dimensional pump, the sample inlet end of the second dimensional chromatographic column, the sample inlet end and the sample outlet end of the trapping column and the ultraviolet detector.
Preferably, the diverter valve is equipped with 1 st interface, 2 nd interface, 3 rd interface, 4 th interface, 5 th interface and 6 th interface at least, first interface with the appearance end intercommunication of catching the post, 2 nd interface with the appearance end intercommunication of second dimension chromatographic column, 3 rd interface with the second dimension pump intercommunication, 4 th interface with the appearance end intercommunication of appearance of catching the post, 5 th interface with the ultraviolet detector intercommunication, 6 th interface with the intercommunication interface intercommunication.
As described above, the present invention has at least one of the following advantageous effects:
1) The method has high sensitivity and good accuracy, and realizes the semi-quantitative determination of 7 HGL-DTGs;
2) The two-dimensional chromatography adopts reversed-phase liquid chromatography, has strong separation capacity, and has the advantages of good compatibility, strong anti-interference capacity, high flux and high peak capacity;
3) The system has the advantages of simple construction, high automation degree and more adjustable parameters;
4) The pretreatment of the analysis method is very simple, the machine can be operated only by extracting and passing a film, and the pretreatment is quick and efficient;
5) The invention establishes an LC multiplied by LC-MS/MS analysis platform based on two-dimensional liquid chromatography, establishes an analysis method for determining tobacco HGL-DTGs by C18 multiplied by C18 two-dimensional liquid chromatography, and realizes qualitative and relative quantitative analysis of tobacco 7 HGL-DTGs.
Drawings
FIG. 1 is a schematic diagram of a two-dimensional liquid chromatography tandem mass spectrometry analysis instrument used in the method for analyzing the content of tobacco hydroxy geranyl linalool diterpene enol glycoside.
FIG. 2 is a process for separating tobacco hydroxy geranyl linalool diterpene enol glycosides HGL-DTGs by two-dimensional liquid chromatography tandem mass spectrometry.
Reference numerals
1. First dimension liquid chromatography
11. First dimension pump
12. First dimension chromatographic column
2. Compensation pump
3. Connection interface
4. Switching valve
41. No. 1 interface
42. 2 nd interface
43. No. 3 interface
44. No. 4 interface
45. 5 th interface
46. 6 th interface
5. Trapping column
6. Second dimension liquid chromatography
61. Second dimension pump
62. Second dimension chromatographic column
7. Mass spectrometry
8. Ultraviolet detector
Detailed Description
The present invention is further illustrated below by reference to specific examples, which are intended to be illustrative only and not to limit the scope of the invention.
A two-dimensional liquid chromatography-mass spectrometry system is shown in figures 1 and 2 and comprises a first-dimensional liquid chromatography 1, a compensation pump 2, a communication interface 3, a switching valve 4, a trapping column 5, a second-dimensional liquid chromatography 6, a mass spectrum 7 and an ultraviolet detector 8;
the first dimension liquid chromatogram 1 comprises a first dimension pump 11 and a first dimension chromatographic column 12 which are communicated, and the second dimension liquid chromatogram 6 comprises a second dimension pump 61 and a second dimension chromatographic column 62 which are communicated;
the communication interface 3 is respectively communicated with the sample outlet end of the first dimension chromatographic column 12, the compensation pump 2 and the switching valve 4 through pipelines;
the sample outlet end of the second-dimension chromatographic column 62 is communicated with the mass spectrum 7;
the switching valve 4 is respectively communicated with the second dimension pump 61, the sample inlet end of the second dimension chromatographic column 62, the sample inlet end and the sample outlet end of the trap column 5 and the ultraviolet detector 8.
In a preferred embodiment, the switching valve 4 is at least provided with a 1 st port 41, a 2 nd port 42, a 3 rd port 43, a 4 th port 44, a 5 th port 45 and a 6 th port 46, the first port 41 is communicated with the sample inlet of the trap column 5, the 2 nd port 42 is communicated with the sample inlet of the second dimension chromatographic column 62, the 3 rd port 43 is communicated with the second dimension pump 61, the 4 th port 44 is communicated with the sample outlet of the trap column 5, the 5 th port 45 is communicated with the ultraviolet detector 8, and the 6 th port 46 is communicated with the communication port 3.
In the invention, the two-dimensional liquid chromatography-mass spectrometry system for analyzing the tobacco hydroxy geranyl linalool diterpene enol glycoside mainly has three stages, as shown in figure 2.
The first stage is first dimension liquid chromatography (reversed phase liquid chromatography) separation, the 1 st interface 41 and the 2 nd interface 42 of the switching valve 4 are connected, the target component HGL-DTGs is separated from impurities, and the impurities directly enter waste liquid;
the second stage is enrichment, the 1 st interface 41 and the 6 th interface 46 of the switching valve 4 are connected, the compensation pump 2 is started, and the target components HGL-DTGs are enriched on the trapping column;
and the third stage is separation and detection of a second-dimension liquid chromatogram (reversed-phase liquid chromatogram), a 1 st interface 41 and a 2 nd interface 42 of the switching valve 4 are connected, target components HGL-DTGs on the trapping column 5 are eluted to the second dimension by 'back flushing', and the target components HGL-DTGs enter a tandem mass spectrum after chromatographic separation, so that qualitative and relative quantitative analysis of 7 types of HGL-DTGs is realized.
1.1 instruments and reagents
Agilent 1260, HPLC; ABI 5500; a green tobacco leaf sample; water, methanol, acetonitrile and formic acid are all chromatographically pure.
1.2 sample pretreatment
Picking green tobacco leaves, transporting by dry ice freezing, pulverizing in liquid nitrogen, grinding, sorting, and storing in a-80 deg.C ultra-low temperature refrigerator for use. A tobacco leaf sample (0.5 g) was weighed, vortexed and shaken for 20 minutes using 10mL of chromatographically pure methanol, centrifuged, and loaded onto a 0.22. Mu. MPTE membrane.
1.3 first dimension chromatography conditions
A chromatographic column: CAPCELL PAK MGIII C18 (50 mm. Times.2.0 mm,3 μm); mobile phase: methanol (plus 0.1% formic acid) -water (plus 0.1% formic acid), flow rate: 0.2mL/min; column temperature: 30 ℃; sample introduction amount: 10.0 μ L; DAD detection wavelength: 210nm, mobile phase gradient as shown in table 1:
TABLE 1 gradient elution procedure for first dimension chromatography mobile phase
1.4 second dimension chromatography conditions
A chromatographic column: agilent ZORBAX 300SB C18 (2.1 mm. Times.100mm, 3.5 μm); mobile phase: acetonitrile (plus 0.1% formic acid) -water (plus 0.1% formic acid), flow rate: 0.4mL/min; column temperature: 30 ℃; the mobile phase gradient is shown in table 2:
TABLE 2 gradient elution procedure for mobile phase of second dimension chromatography
1.5 trapping conditions
A trapping column: thermo scientific Acclaim Polar Advantage II (4.6 mm. Times.10mm, 5 μm); fluid infusion: water; column temperature: 30 ℃; cutting time: 16.8min-21.8min; the flow rate of the liquid supplement is 0-15min, and the flow rate is 0.1mL/min;16-22min,1.0mL/min;23-30min,0.1mL/min.
1.6 Mass Spectrometry parameter conditions
Electrospray ion source (ESI), negative ion scanning, multiple reaction detection (MRM) mode; electrospray voltage: -4500V; ion source temperature: 500 ℃; residence monitoring time of ion pair (dwell time): 20ms; atomizing gas pressure: 50Psi; auxiliary gas pressure: 50Psi; mass spectrometry scan time: 27-37min. The other main detection parameters of the mass spectrum of 7 HGL-DTG compounds are shown in Table 3:
TABLE 3 tandem Mass Spectrometry ion Pair parameters
2. Reproducibility of the method
A variety of samples of green tobacco leaves were selected and the reproducibility of the method was examined for 7 different HGL-DTGs compounds (n = 3) and the results are shown in Table 4. The repeatability RSD of the sample is in the range of 0.24-7.21%, and the repeatability is better.
TABLE 4 in-day reproducibility of HGL-DTGs (n = 3)
3. Analysis of actual samples
The relative contents of HGL-DTGs of 10 different green tobacco leaf samples are detected by using a 2DLC-MS method, and the peak area results of the contents of HGL-DTGs are shown in a table 5.
TABLE 5 Green tobacco leaf sample HGL-DTGs content results
The above examples are intended to illustrate the disclosed embodiments of the invention and are not to be construed as limiting the invention. In addition, various modifications of the methods and compositions set forth herein, as well as variations of the methods and compositions of the present invention, will be apparent to those skilled in the art without departing from the scope and spirit of the invention. While the invention has been specifically described in connection with various specific preferred embodiments thereof, it should be understood that the invention is not limited to those specific embodiments. Indeed, various modifications of the above-described embodiments which are obvious to those skilled in the art to which the invention pertains are intended to be covered by the scope of the present invention.
Claims (10)
1. An analysis method of tobacco hydroxy geranyl linalool diterpene enol glycoside is characterized by comprising the following steps: pretreating tobacco leaves, and qualitatively and quantitatively analyzing samples to be detected obtained by pretreatment by adopting a two-dimensional liquid chromatography-mass spectrometry combined method: separating out a target component through a first-dimension liquid chromatogram, trapping the target component after dilution by a solution supplement, eluting the trapped component, separating through a second-dimension liquid chromatogram, and performing qualitative and quantitative analysis through a mass spectrum.
2. The method of analyzing tobacco hydroxy-geranyl linalool diterpenenol glycosides according to claim 1, wherein the pre-treatment comprises the steps of:
1) Crushing and grinding the tobacco leaves under liquid nitrogen;
2) Oscillating the tobacco leaves obtained in the step 1) in an organic solvent, centrifuging and then coating a film to obtain a sample to be detected.
3. The method of claim 1, further comprising at least one of the following technical features:
a) The conditions of the first dimension liquid chromatogram are as follows: first dimension liquid chromatography: reversed phase liquid chromatography; a first dimension chromatographic column: CAPCELL PAK MGIIIC18; flow rate: 0.05-1 mL/min; column temperature: 20 to 60 ℃; sample introduction amount: 1-20.0 mu L; first dimension mobile phase a phase: water plus 0.05 to 2 percent of formic acid; first dimension mobile phase B phase: methanol and 0.05 to 2 percent of formic acid; gradient leaching;
b) The conditions of the second dimension liquid chromatogram are as follows: second dimension liquid chromatography: reversed phase liquid chromatography; a second dimension chromatographic column: agilent ZORBAX 300SB C18; flow rate: 0.05-1 mL/min; column temperature: 20 to 60 ℃; second-dimensional mobile phase a phase: water and 0.05 to 2 percent of formic acid; second dimension mobile phase B phase: acetonitrile and 0.05-2% formic acid; and (4) gradient elution.
4. The method for analyzing tobacco hydroxy geranyl linalool diterpenenol glycoside according to claim 3, wherein in characteristic a), the specific procedure of gradient elution is as follows:
0-5 min, first dimension mobile phase A phase: the volume ratio of the first-dimension mobile phase B phase is 70:30;
5-20 min, first dimension mobile phase A phase: the volume ratio of the first-dimension mobile phase B phase is 70: 30-5: 95;
20-25 min, first dimension mobile phase A phase: the volume ratio of the first-dimension mobile phase B phase is 5:95;
25-26 min, first dimension mobile phase A phase: the volume ratio of the first-dimensional mobile phase B phase is 5: 95-70: 30, of a nitrogen-containing gas;
26-50 min, first dimension mobile phase A phase: the volume ratio of the first-dimension mobile phase B phase is 70:30.
5. the method for analyzing tobacco hydroxy geranyl linalool diterpenenol glycoside according to claim 3, wherein in characteristic b), the specific procedure of the gradient elution is:
0-22 min, second dimension mobile phase A phase: the volume ratio of the second-dimensional mobile phase B phase is 95:5;
22-27 min, second dimension mobile phase A phase: the volume ratio of the second-dimensional mobile phase B phase is 95:5 to 70:30, of a nitrogen-containing gas;
27-37min, second dimension mobile phase A phase: the volume ratio of the second-dimension mobile phase B phase is 70:30 to 40:60, adding a solvent to the mixture;
37-37.1 min, second dimension mobile phase A phase: the volume ratio of the second-dimension mobile phase B phase is 40: 60-5: 95;
37.1-45 min, second dimension mobile phase A phase: the volume ratio of the second-dimension mobile phase B phase is 5:95;
45-45.1 min, second dimension mobile phase A phase: the volume ratio of the second-dimension mobile phase B phase is 5: 95-95: 5;
45.1-50 min, second dimension mobile phase A phase: the volume ratio of the second-dimensional mobile phase B phase is 95:5.
6. the method for analyzing tobacco hydroxy geranyl linalool diterpenenol glycoside according to claim 1, wherein the trapping conditions are:
a trapping column: thermo scientific Acclaim Polar Advantage II;
liquid supplementing: water;
column temperature: 20 to 60 ℃;
cutting time: 16.8min-21.8min;
the flow rate of the liquid supplement is 0-15min, and the flow rate is 0.1-1 mL/min;16-22min, 0.1-1.0 mL/min;23 to 30min,0.1 to 1mL/min.
7. The method for analyzing tobacco hydroxy geranyl linalool diterpenenol glycoside according to claim 1, wherein the conditions of the mass spectrometry are: electrospray ion source, negative ion scanning and multi-reaction detection MRM mode; electrospray voltage: -3000-6000V; ion source temperature: 200 to 600 ℃; residence monitoring time of ion pair: 10-100 ms; atomizing gas pressure: 20 to 60Psi; auxiliary gas pressure: 20 to 60Psi; mass spectrometry scan time: 27-37min.
8. The method of claim 1, wherein the tobacco hydroxy geranyl linalool diterpenol glycoside comprises lycoumoside I, nicotianoside III, nicotianoside IV, nicotianoside V, nicotianoside VI, and Nicotianoside VII.
9. A two-dimensional liquid chromatography-mass spectrometry combined system is characterized by comprising a first-dimensional liquid chromatography (1), a compensation pump (2), a communication interface (3), a switching valve (4), a trapping column (5), a second-dimensional liquid chromatography (6), a mass spectrum (7) and an ultraviolet detector (8);
the first dimension liquid chromatogram (1) comprises a first dimension pump (11) and a first dimension chromatographic column (12) which are communicated, and the second dimension liquid chromatogram (6) comprises a second dimension pump (61) and a second dimension chromatographic column (62) which are communicated;
the communication interface (3) is respectively communicated with the sample outlet end of the first dimension chromatographic column (12), the compensation pump (2) and the switching valve (4) through pipelines;
the sample outlet end of the second-dimension chromatographic column (62) is communicated with the mass spectrum (7);
the switching valve (4) is respectively communicated with the second dimensional pump (61), the sample introduction end of the second dimensional chromatographic column (62), the sample introduction end and the sample outlet end of the trapping column (5) and the ultraviolet detector (8).
10. The two-dimensional liquid chromatography-mass spectrometry system of claim 9, wherein the switching valve (4) is provided with at least a 1 st port (41), a 2 nd port (42), a 3 rd port (43), a 4 th port (44), a 5 th port (45) and a 6 th port (46), the first port (41) is communicated with a sample inlet of the trap column (5), the 2 nd port (42) is communicated with a sample inlet of the second dimension chromatographic column (62), the 3 rd port (43) is communicated with the second dimension pump (61), the 4 th port (44) is communicated with a sample outlet of the trap column (5), the 5 th port (45) is communicated with the ultraviolet detector (8), and the 6 th port (46) is communicated with the communication port (3).
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CN106053675A (en) * | 2016-08-02 | 2016-10-26 | 上海烟草集团有限责任公司 | Method for analyzing nitrosamine release amount in cigarette smoke through double-column liquid chromatogram tandem mass spectrometry |
CN110632220A (en) * | 2019-10-31 | 2019-12-31 | 上海烟草集团有限责任公司 | Method for analyzing sucrose ester in tobacco by multi-dimensional liquid chromatography-mass spectrometry |
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CN106053675A (en) * | 2016-08-02 | 2016-10-26 | 上海烟草集团有限责任公司 | Method for analyzing nitrosamine release amount in cigarette smoke through double-column liquid chromatogram tandem mass spectrometry |
CN110632220A (en) * | 2019-10-31 | 2019-12-31 | 上海烟草集团有限责任公司 | Method for analyzing sucrose ester in tobacco by multi-dimensional liquid chromatography-mass spectrometry |
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