Method for detecting carotenoid in sorghum based on liquid chromatography-mass spectrometry

Abstract

The invention relates to the technical field of wine brewing, discloses a method for detecting carotenoid in sorghum based on liquid chromatography-mass spectrometry, and aims to solve the problem that the existing carotenoid detection mode is poor in efficiency and accuracy, and the scheme mainly comprises the following steps: preparing a sample solution of sorghum to be detected; obtaining standard solutions of a plurality of carotenoid standard substances; respectively carrying out liquid chromatography-mass spectrometry analysis and detection on the sample solution and the standard solution in a full scanning mode; obtaining a carotenoid qualitative detection result of the sample solution according to analysis and detection of the sample solution and the standard solution; preparing a reference substance solution of carotenoid standard substances, selecting an ion monitoring mode to scan the reference substance solution, and drawing a standard curve corresponding to each carotenoid standard substance; and calculating the content of each carotenoid in the sample solution. The carotenoid detection efficiency and accuracy are improved, and the method is particularly suitable for brewing sorghum.

Description

Technical Field

[0001] The present application relates to the field of wine brewing technology, and in particular to a method for detecting carotenoids in sorghum based on liquid chromatography-mass spectrometry. Background Art

[0002] There are huge sensory differences between liquors made from different grains. In the brewing industry, sorghum plays a vital role. As the main raw material for brewing liquor, the quality of sorghum directly determines the quality of the liquor. Its starch, protein, fat, tannin and other nutrients are degraded and synthesized by microorganisms to form ethanol and a series of aromatic and flavorful substances, which together create the body style of the liquor.

[0003] Carotenoids are a general term for an important class of natural pigments. They are a class of terpenoid substances with multiple conjugated double bonds that are yellow, orange or red in color. Carotenoids are a class of natural pigments with antioxidant and anti-inflammatory properties found in photosynthetic organisms. They are a class of pigments that cannot be synthesized by humans and animals themselves and must be taken from the outside world. They are also the main source of vitamin A in the body. There are many types of carotenoids, and different carotenoids have different physiological functions and a wide range of applications. In recent years, the biological functions provided by carotenoids in grains have gradually attracted attention, and the content and type of carotenoids are becoming an important quality feature of grains.

[0004] At present, there are many methods for analyzing and detecting carotenoids, including ultraviolet spectrophotometry (UV), thin layer chromatography (TLC), high performance liquid chromatography (HPLC), ultra high performance liquid chromatography (UPLC), liquid chromatography-triple quadrupole mass spectrometry (LC-MS / MS), etc. However, these methods require the optimization of product ions and related parameters for the target objects one by one. Not only does it take a long time to detect, it is also easily interfered by the matrix and produces false positives, and has poor efficiency and accuracy. Summary of the invention

[0005] The present application aims to solve the problem of poor efficiency and accuracy of existing methods for detecting carotenoids, and proposes a method for detecting carotenoids in sorghum based on liquid chromatography-mass spectrometry.

[0006] The technical solution adopted by this application to solve the above technical problems is:

[0007] A method for detecting carotenoids in sorghum based on liquid chromatography-mass spectrometry, the method comprising:

[0008] Prepare sample solutions of sorghum to be tested and obtain standard solutions of multiple carotenoid standards;

[0009] Based on the set liquid phase conditions and mass spectrometry conditions, the sample solution and the standard solution are respectively subjected to liquid chromatography-mass spectrometry analysis and detection in full scan mode;

[0010] Establishing a carotenoid database according to the analysis and detection results of the standard solution, comparing the analysis and detection results of the sample solution with the carotenoid database, and obtaining a qualitative detection result of carotenoids of the sample solution;

[0011] Prepare a reference solution of carotenoid standards, select an ion monitoring mode to scan the reference solution, and obtain mass spectrometry peak intensity data of quantitative signals of each carotenoid standard in the reference solution;

[0012] According to the mass spectrometry peak intensity data, a standard curve corresponding to each carotenoid standard is plotted with the standard concentration as the abscissa and the peak area of ​​the mass spectrometry peak as the ordinate, and a linear equation corresponding to each standard curve is determined;

[0013] The contents of various types of carotenoids in the sample solution are calculated according to the qualitative detection results of carotenoids in the sample solution and the corresponding linear equations to obtain the quantitative detection results of carotenoids in the sample solution.

[0014] Further, preparing a sample solution of sorghum to be tested specifically comprises:

[0015] Weigh sorghum powder, add extract solution, mix and shake well, the extract solution is hexane, acetone and ethanol containing 0.01% BHT, and the ratio of hexane, acetone and ethanol is 2:1:1;

[0016] Low-temperature ultrasonic extraction for 1 h, centrifugation at 8000 rpm for 8 min at 4 °C, collection of the supernatant, and repeated three to four times until the extract is colorless;

[0017] Add saturated saline solution to the collected supernatant, shake vigorously for 30 seconds, let stand to separate the layers, collect the upper layer, blow with concentrated nitrogen to obtain the sample solution.

[0018] Furthermore, the liquid phase conditions are:

[0019] Chromatographic column: YMC C30 Carotenoid (150mm*2.1*2.6μm);

[0020] Mobile phase A: methanol: acetonitrile: water, mobile phase B: methyl tert-butyl ether;

[0021] Column temperature: 20°C;

[0022] Injection volume: 1ul;

[0023] Flow rate: 0.2ml / min;

[0024] Elution gradient program:

[0025] 0min-2min, the volume ratio of mobile phase A to mobile phase B was 100:0;

[0026] 15 min, the volume ratio of mobile phase A to mobile phase B was 60:40;

[0027] 18 min, the volume ratio of mobile phase A to mobile phase B was 20:80;

[0028] 22min, the volume ratio of mobile phase A to mobile phase B was 0:100.

[0029] Furthermore, the mass spectrometry conditions are:

[0030] Scan mode: Full scan-SIM;

[0031] Ion source: APCI+;

[0032] Ion transfer tube temperature: 300°C;

[0033] Ion source heating temperature: 350℃;

[0034] Auxiliary gas: 5Arb;

[0035] Sheath gas: 45Arb;

[0036] Primary resolution: 12000;

[0037] SIM mode resolution: 60000;

[0038] Scan range: 40-1400.

[0039] Further, the plurality of carotenoid standards include neoxanthin, antheraxanthin, 3′-epi-lutein, lutein, capsanthin, zeaxanthin, E / Z-phytoene, echinenone, E / Z-phytoene and β-carotene.

[0040] Further, the carotenoid qualitative detection result of the sample solution is obtained, specifically including:

[0041] Data screening is performed on the analysis and detection results of the sample solution, and the retention time and secondary fragment ion information of each carotenoid in the screening results are compared with the carotenoid database to obtain the qualitative detection result of the carotenoids in the sample solution.

[0042] Further, obtaining the quantitative detection result of carotenoids in the sample solution specifically includes:

[0043] The integrated peak area ratio of carotenoids in the analysis and detection results of the sample solution is substituted into the corresponding standard equation to calculate the concentration, and the content of the corresponding carotenoid in the sample solution is calculated according to the concentration.

[0044] Furthermore, the calculation formula for the content of the corresponding carotenoids in the sample solution is as follows:

[0045]

[0046] Wherein, M represents the content of the corresponding carotenoid in the sample solution, c represents the concentration of the corresponding carotenoid in the sample solution, V represents the volume of the sample solution, and m represents the mass of the sample solution.

[0047] Furthermore, the liquid chromatography-mass spectrometry is a Vanquish Flex Binary ultra-high performance liquid chromatograph connected in series with an Orbitrap Exploris 120 mass spectrometer.

[0048] The beneficial effects of the present application are as follows: the method for detecting carotenoids in sorghum based on liquid chromatography-mass spectrometry provided in the present application performs qualitative analysis in full scan mode by an ultra-high performance liquid chromatography-mass spectrometer, and then directly quantifies the target substance by using the precise mass-to-charge ratio of the target substance parent ion in selected ion monitoring mode, without the need to optimize the daughter ions and related parameters of the target substance one by one, thereby reducing the detection time and effectively avoiding false positive phenomena caused by matrix interference, thereby improving the detection efficiency and accuracy, and by establishing a carotenoid standard database, it is possible to more accurately analyze and detect related carotenoids. BRIEF DESCRIPTION OF THE DRAWINGS

[0049] Figure 1 A schematic diagram of a method for detecting carotenoids in sorghum based on liquid chromatography-mass spectrometry provided in an embodiment of the present application;

[0050] Figure 2 A standard spectrum corresponding to lutein provided in an embodiment of the present application;

[0051] Figure 3 A standard spectrum corresponding to zeaxanthin provided in the embodiment of the present application;

[0052] Figure 4 A standard spectrum corresponding to anther xanthin provided in the embodiment of the present application;

[0053] Figure 5 A total ion current diagram of a sample solution provided in an embodiment of the present application;

[0054] Figure 6 A total ion current diagram of a reference solution provided in an embodiment of the present application;

[0055] Figure 7 A schematic diagram of a standard curve of echinenone provided in an embodiment of the present application;

[0056] Figure 8 A schematic diagram of a standard curve of E / Z-phytoene provided in an embodiment of the present application;

[0057] Fig. 9 A schematic diagram of a standard curve of E / Z-phytoene provided in an embodiment of the present application. DETAILED DESCRIPTION

[0058] In order to enable those skilled in the art to better understand the solution of the present application, the technical solution in the embodiments of the present application will be clearly and completely described below in conjunction with the drawings in the embodiments of the present application.

[0059] The technical solution of the embodiment of the present application is applicable to application scenarios where it is necessary to detect the type and content of carotenoids in sorghum.

[0060] Since the current carotenoid detection method requires optimizing the daughter ions and related parameters for each target, not only is the detection time long, but it is also easily affected by matrix interference and produces false positives, and the efficiency and accuracy are poor. Based on this, the technical scheme of the present application is proposed. In an embodiment of the present application, a sample solution of sorghum to be tested is prepared, and a plurality of standard solutions of carotenoid standards are obtained; based on the set liquid phase conditions and mass spectrometry conditions, the sample solution and the standard solution are respectively subjected to liquid chromatography-mass spectrometry analysis and detection in full scan mode; a carotenoid database is established according to the analysis and detection results of the standard solution, and the analysis and detection results of the sample solution are compared with the carotenoid database to obtain a qualitative detection result of carotenoids in the sample solution; a reference solution of carotenoid standards is prepared, and the reference solution is scanned in a selected ion monitoring mode to obtain mass spectrometry peak intensity data of quantitative signals of each carotenoid standard in the reference solution; based on the mass spectrometry peak intensity data, a standard curve corresponding to each carotenoid standard is plotted with the standard concentration as the horizontal coordinate and the peak area of ​​the mass spectrometry peak as the vertical coordinate, and a linear equation corresponding to each standard curve is determined; the content of each carotenoid in the sample solution is calculated according to the qualitative detection result of carotenoids in the sample solution and the corresponding linear equation to obtain a quantitative detection result of carotenoids in the sample solution.

[0061] Specifically, the embodiment of the present application performs analysis and detection by ultra-high performance liquid chromatography-mass spectrometry, analyzes and detects the sample solution and the standard solution in full scan mode, and establishes a carotenoid standard database using the analysis and detection results of the standard solution, thereby confirming the carotenoids contained in the sample solution according to the analysis and detection results of the sample solution, and then prepares a reference solution using relevant carotenoid standards, and scans it in ion monitoring mode to establish a standard curve, and uses this to detect the carotenoid content in the sample solution, specifically using the precise mass-to-charge ratio of the target parent ion for direct quantification, thereby eliminating the need to optimize the daughter ions and related parameters of the target one by one, reducing the detection time while avoiding false positives caused by matrix interference, thereby improving detection efficiency and accuracy.

[0062] The technical solutions in the embodiments of the present application will be clearly and completely described below in conjunction with the drawings in the embodiments of the present application. Obviously, the described embodiments are only part of the embodiments of the present application, rather than all the embodiments.

[0063] See also Figure 1 The method for detecting carotenoids in sorghum based on liquid chromatography-mass spectrometry provided in the embodiment of the present application comprises the following steps:

[0064] S1. Prepare a sample solution of sorghum to be tested, and obtain standard solutions of multiple carotenoid standards.

[0065] In the embodiment of the present application, the method for preparing the sample solution of sorghum to be detected specifically comprises the following steps:

[0066] S11, weighing sorghum powder, adding extracting solution, mixing and shaking, wherein the extracting solution is hexane, acetone and ethanol containing 0.01% BHT, and the ratio of hexane, acetone and ethanol is 2:1:1;

[0067] Among them, the sorghum can be ground into powder under low temperature and light-proof conditions, and the sorghum powder can be stored in a -40°C refrigerator for later use.

[0068] S12, low temperature ultrasonic extraction for 1 hour, centrifugation at 8000 rpm for 8 minutes at 4°C, collecting the supernatant, repeating the operation three to four times until the extract is colorless;

[0069] S13. Add saturated saline solution to the collected supernatant, shake vigorously for 30 seconds, let stand to separate the layers, collect the upper layer, blow with concentrated nitrogen to obtain the sample solution.

[0070] During the whole process of preparing the sample solution of sorghum to be tested, a low temperature and light-proof environment was maintained.

[0071] In the examples of the present application, carotenoid standards include neoxanthin, antheraxanthin, 3'-epi-lutein, lutein, capsanthin, zeaxanthin, E / Z-phytoene, echinenone, E / Z-phytoene and β-carotene. The carotenoid standards are dissolved in methyl tert-butyl ether containing BHT and stored under argon.

[0072] S2. Based on the set liquid phase conditions and mass spectrometry conditions, the sample solution and the standard solution are respectively subjected to liquid chromatography-mass spectrometry analysis and detection in full scan mode.

[0073] In the embodiments of the present application, an ultra-high performance liquid chromatography-mass spectrometry (UPLC-MS) is used to detect carotenoids. The core of UPLC-MS technology is to use ultra-high performance liquid chromatography as the injection system of mass spectrometry, so that complex chemical components can be effectively separated, and mass spectrometry is used as a detector for quantitative and qualitative analysis. In practical applications, UPLC-MS can use a Vanquish Flex Binary ultra-high performance liquid chromatograph in series with an Orbitrap Exploris 120 mass spectrometer.

[0074] In the examples of the present application, the liquid phase conditions are:

[0075] Chromatographic column: YMC C30 Carotenoid (150mm*2.1*2.6μm);

[0076] Mobile phase A: methanol: acetonitrile: water, mobile phase B: methyl tert-butyl ether;

[0077] Column temperature: 20°C;

[0078] Injection volume: 1ul;

[0079] Flow rate: 0.2ml / min;

[0080] Elution gradient program:

[0081] 0min-2min, the volume ratio of mobile phase A to mobile phase B was 100:0;

[0082] 15 min, the volume ratio of mobile phase A to mobile phase B was 60:40;

[0083] 18 min, the volume ratio of mobile phase A to mobile phase B was 20:80;

[0084] 22min, the volume ratio of mobile phase A to mobile phase B was 0:100.

[0085] The mass spectrometry conditions are:

[0086] Scan mode: Full scan-SIM;

[0087] Ion source: APCI+;

[0088] Ion transfer tube temperature: 300°C;

[0089] Ion source heating temperature: 350℃;

[0090] Auxiliary gas: 5Arb;

[0091] Sheath gas: 45Arb;

[0092] Primary resolution: 12000;

[0093] SIM mode resolution: 60000;

[0094] Scan range: 40-1400.

[0095] In practical applications, each standard solution is analyzed and tested on the machine according to the above liquid phase conditions and mass spectrometry conditions, the retention time and characteristic ion information of the carotenoids corresponding to each standard solution are determined, the analysis and detection results of the standard solution are obtained, and the corresponding standard spectrum is generated. Figures 2 to 4 The standard spectra corresponding to lutein, zeaxanthin and antheraxanthin are schematically shown, with the abscissa being the mass-to-charge ratio and the ordinate being the ion intensity.

[0096] Similarly, the sample solution is analyzed and tested on the machine according to the above liquid phase conditions and mass spectrometry conditions to determine the retention time and characteristic ion information of each carotenoid in the sample solution, and obtain the analysis and detection results of the sample solution. Specifically, when the components flow into the mass spectrometer after chromatographic separation, the mass spectrometer continuously scans for data acquisition, and each scan obtains a mass spectrum. All ion intensities in each mass spectrum are added together to obtain a total ion current intensity, and a corresponding total ion current diagram is generated. Please refer to Figure 5 , the horizontal axis is time and the vertical axis is the total ion current intensity.

[0097] S3. Establishing a carotenoid database according to the analysis and detection results of the standard solution, comparing the analysis and detection results of the sample solution with the carotenoid database, and obtaining a qualitative detection result of carotenoids of the sample solution.

[0098] In practical applications, the analysis and detection results of the sample solution are first screened, and the retention time and secondary fragment ion information of each carotenoid in the screening results are compared with the carotenoid database to obtain the qualitative detection results of the carotenoids in the sample solution. For example, the carotenoids contained in the sample solution are identified to include antheraxanthin, lutein, zeaxanthin, E / Z-phytoene, echinenone, and E / Z-phytoene.

[0099] S4. Prepare a reference solution of carotenoid standards according to the carotenoid qualitative detection result, select an ion monitoring mode to scan the reference solution, and obtain mass spectrum peak intensity data of the quantitative signal of each carotenoid standard in the reference solution.

[0100] Specifically, after completing the qualitative detection of the sample solution, prepare the reference solution of each carotenoid standard, then set the mass-to-charge ratio and scanning time range for the acquisition, select the ion monitoring mode to scan the reference solution, obtain the total ion current diagram of the reference solution in SIM mode, and then obtain the mass spectrum peak intensity data of the corresponding quantitative signal of each carotenoid standard. Please refer to Figure 6 , the horizontal axis is time and the vertical axis is the total ion current intensity.

[0101] S5. According to the mass spectrum peak intensity data, a standard curve corresponding to each carotenoid standard is drawn with the standard concentration as the abscissa and the peak area of ​​the mass spectrum peak as the ordinate, and a linear equation corresponding to each standard curve is determined.

[0102] Figures 7 to 9 The standard curves corresponding to echinenone, E / Z-phytoene and E / Z-phytoene are schematically shown, and the corresponding linear equations are: Y=102838e+007*X; Y=-1.49881+302473*X; Y=-36262.3+800988*X, wherein X represents the concentration of the standard and Y represents the peak area of ​​the mass spectrum peak.

[0103] S6. Calculate the content of each type of carotenoid in the sample solution according to the qualitative detection result of carotenoids in the sample solution and the corresponding linear equation to obtain the quantitative detection result of carotenoids in the sample solution.

[0104] Specifically, in the analysis and detection results of the sample solution, the integrated peak area ratio corresponding to the detected carotenoids is substituted into the corresponding standard equation to calculate the concentration, and the content of the corresponding carotenoids in the sample solution is calculated according to the concentration to complete the quantitative detection of the carotenoids in the sample solution. The calculation formula is as follows:

[0105]

[0106] Wherein, M represents the content of the corresponding carotenoid in the sample solution, c represents the concentration of the corresponding carotenoid in the sample solution, V represents the volume of the sample solution, and m represents the mass of the sample solution.

[0107] In summary, the method for detecting carotenoids in sorghum based on liquid chromatography-mass spectrometry provided in the embodiment of the present application performs qualitative analysis in full scan mode by ultra-high performance liquid chromatography-mass spectrometry, and then directly quantifies by using the precise mass-to-charge ratio of the target parent ion in selected ion monitoring mode, without the need to optimize the daughter ions and related parameters of the target one by one, thereby reducing the detection time and avoiding false positives caused by matrix interference, thereby improving the detection efficiency and accuracy, and by establishing a carotenoid standard database, it is possible to more accurately analyze and detect related carotenoids. This detection method can quickly and accurately quantify carotenoids in different special grains for brewing, with the quantitative limits of each compound being less than 2.4 ppm and the linear correlation coefficient R 2 All of them were greater than 0.997, and the relative standard deviation was less than 5%, which provided a new detection standard for exploring carotenoids in grains and was of great significance for improving the quality of sorghum.

Claims

1. A method for detecting carotenoids in sorghum based on liquid chromatography-mass spectrometry, characterized in that: The method comprises: Prepare sample solutions of sorghum to be tested and obtain standard solutions of multiple carotenoid standards; Based on the set liquid phase conditions and mass spectrometry conditions, the sample solution and the standard solution are respectively subjected to liquid chromatography-mass spectrometry analysis and detection in full scan mode; Establishing a carotenoid database according to the analysis and detection results of the standard solution, comparing the analysis and detection results of the sample solution with the carotenoid database, and obtaining a qualitative detection result of carotenoids of the sample solution; Prepare a reference solution of carotenoid standards, select an ion monitoring mode to scan the reference solution, and obtain mass spectrometry peak intensity data of quantitative signals of each carotenoid standard in the reference solution; According to the mass spectrometry peak intensity data, a standard curve corresponding to each carotenoid standard is plotted with the standard concentration as the abscissa and the peak area of ​​the mass spectrometry peak as the ordinate, and a linear equation corresponding to each standard curve is determined; The contents of various types of carotenoids in the sample solution are calculated according to the qualitative detection results of carotenoids in the sample solution and the corresponding linear equations to obtain the quantitative detection results of carotenoids in the sample solution.

2. The method for detecting carotenoids in sorghum based on liquid chromatography-mass spectrometry according to claim 1, characterized in that: Prepare the sample solution of sorghum to be tested, including: Weigh sorghum powder, add extract solution, mix and shake well, the extract solution is hexane, acetone and ethanol containing 0.01% BHT, and the ratio of hexane, acetone and ethanol is 2:1:1; Low-temperature ultrasonic extraction for 1 h, centrifugation at 8000 rpm for 8 min at 4 °C, collection of the supernatant, and repeated three to four times until the extract is colorless; Add saturated saline solution to the collected supernatant, shake vigorously for 30 seconds, let stand to separate the layers, collect the upper layer, blow with concentrated nitrogen to obtain the sample solution.

3. The method for detecting carotenoids in sorghum based on liquid chromatography-mass spectrometry according to claim 1, characterized in that: The liquid phase conditions are: Chromatographic column: YMC C30 Carotenoid (150mm*2.1*2.6μm); Mobile phase A: methanol: acetonitrile: water, mobile phase B: methyl tert-butyl ether; Column temperature: 20°C; Injection volume: 1ul; Flow rate: 0.2ml / min; Elution gradient program: 0min-2min, the volume ratio of mobile phase A to mobile phase B was 100:0; 15 min, the volume ratio of mobile phase A to mobile phase B was 60:40; 18 min, the volume ratio of mobile phase A to mobile phase B was 20:80; 22min, the volume ratio of mobile phase A to mobile phase B was 0:

100.

4. The method for detecting carotenoids in sorghum based on liquid chromatography-mass spectrometry according to claim 1, characterized in that: The mass spectrometry conditions are: Scan mode: Full scan-SIM; Ion source: APCI+; Ion transfer tube temperature: 300°C; Ion source heating temperature: 350℃; Auxiliary gas: 5Arb; Sheath gas: 45Arb; Primary resolution: 12000; SIM mode resolution: 60000; Scan range: 40-1400.

5. The method for detecting carotenoids in sorghum based on liquid chromatography-mass spectrometry according to claim 1, characterized in that: The plurality of carotenoid standards include neoxanthin, antheraxanthin, 3′-epi-lutein, lutein, capsanthin, zeaxanthin, E / Z-phytoene, echinenone, E / Z-phytoene and β-carotene.

6. The method for detecting carotenoids in sorghum based on liquid chromatography-mass spectrometry according to claim 1, characterized in that: The carotenoid qualitative test results of the sample solution are obtained, including: Data screening is performed on the analysis and detection results of the sample solution, and the retention time and secondary fragment ion information of each carotenoid in the screening results are compared with the carotenoid database to obtain the qualitative detection result of the carotenoids in the sample solution.

7. The method for detecting carotenoids in sorghum based on liquid chromatography-mass spectrometry according to claim 1, characterized in that: Obtaining the quantitative detection results of carotenoids in the sample solution, specifically including: The integrated peak area ratio of carotenoids in the analysis and detection results of the sample solution is substituted into the corresponding standard equation to calculate the concentration, and the content of the corresponding carotenoid in the sample solution is calculated according to the concentration.

8. The method for detecting carotenoids in sorghum based on liquid chromatography-mass spectrometry according to claim 7, characterized in that: The calculation formula for the content of the corresponding carotenoids in the sample solution is as follows: Wherein, M represents the content of the corresponding carotenoid in the sample solution, c represents the concentration of the corresponding carotenoid in the sample solution, V represents the volume of the sample solution, and m represents the mass of the sample solution.

9. The method for detecting carotenoids in sorghum based on liquid chromatography-mass spectrometry according to any one of claims 1 to 8, characterized in that: The liquid chromatography-mass spectrometry is a Vanquish Flex Binary ultra-high performance liquid chromatograph connected in series with an Orbitrap Exploris 120 mass spectrometer.

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