JP4335106B2 - Mass spectrometer - Google Patents

Description

  The present invention relates to a mass spectrometer, and more particularly to sample ionization in a mass spectrometer.

  In a mass spectrometer that performs mass analysis of ions derived from a sample, there is an MS / MS method as a method for acquiring structural information of sample molecules. In this MS / MS method, sample molecule ions are introduced into a collision cell containing a collision gas such as helium, and sample molecules are cleaved by a collision-induced cleavage reaction between the collision gas and sample molecule ions. The mass spectrum of the product ion is acquired and the structure is analyzed.

  At this time, the reaction efficiency of the molecular ion to be analyzed varies depending on the molecular weight, the type of functional group, the molecular structure, and the like.

  In the MS / MS method, it is necessary to adjust the conditions of the cleavage reaction. However, every time the cleavage conditions are changed, an adjustment operation is required in which a sample is introduced and the conditions are determined from the obtained results.

  Therefore, Patent Document 1 discloses a technique capable of executing the condition adjustment of the cleavage reaction by one sample injection.

  The technique disclosed in Patent Document 1 is to obtain mass spectrum data by causing a specific ion to undergo a cleavage reaction while sweeping a cleavage condition with a single component a plurality of times while sweeping the mass number.

  Here, a three-dimensional quadrupole mass spectrometer (hereinafter abbreviated as 3DQMS) is known as a mass spectrometer. In this 3DQMS, a ring electrode is arranged in an insulator for keeping and fixing the 3DQ part, and end cap electrodes are arranged above and below the insulator to constitute a 3DQQ part.

  The sample molecule ions ionized inside or outside the 3DQ and introduced into the 3DQ are held in the 3DQ part for an arbitrary time, and then released from the pores of the end cap electrode to the detector for each mass number.

  In order to discharge ions accumulated in the 3DQ within an arbitrary time to the outside of the 3DQ for each mass number and introduce them to the detector, a high-frequency voltage corresponding to the ion mass number is continuously swept through the mass range for a certain period of time. The ion is discharged for every mass number.

  When analysis is performed by the above-described 3DQMS by the MS / MS method, collision gas such as helium and ions derived from sample molecules are introduced into 3DQ and reacted.

JP 2000-171442 A

  By the way, in a mass spectrometer, when analyzing multiple components simultaneously by MS / MS method, there is a difference in the cleavage efficiency of each component, and the analysis may be difficult under the same cleavage conditions.

  For this reason, in the prior art, it was necessary to perform analysis a plurality of times by operating conditions such as the gas type and gas flow rate for each analysis.

  In the technique described in Patent Document 1 described above, the condition adjustment of the cleavage reaction can be performed by one sample injection. When changing the cleavage conditions, the gas type and the gas flow rate are changed for each change. It was necessary to manipulate and set conditions such as the above, and it was necessary to perform many measurements.

  In addition, in order to execute MS / MS methods by multiple methods during one analysis using a single device, the flow path of molecular ions is branched and introduced into multiple collision cells, and each is cleaved under different conditions. However, there are problems such as an increase in the size of the apparatus, complicated operation, and high costs.

  An object of the present invention is to realize a mass spectrometer capable of acquiring detailed structural information of a component to be analyzed while suppressing an increase in size of the apparatus and shortening a measurement time.

  In order to achieve the above object, the present invention is configured as follows.

  (1) The mass spectrometer of the present invention has a collision cell (3DQ portion) for performing a collision-induced cleavage reaction between ions derived from a sample and a collision gas, and performs mass analysis of the ions.

The mass spectrometer includes a gas introduction path for introducing a plurality of types of collision gases into the collision cell alone or in combination, and a flow rate for introducing the plurality of types of collision gases into the collision cell. Gas flow rate adjusting means for adjusting each of the collision gas, and control for controlling the gas flow rate adjusting means to adjust the time variation of the type, flow rate, and collision gas mixing ratio of the collision gas introduced into the collision cell. Means, an exhaust port formed in the collision cell for adjusting the pressure inside the collision cell, and a control valve for controlling the amount of exhaust gas from the exhaust port .

(2) Preferably, in (1) above, the control means controls the electric field applied to the collision cell, and the voltage value applied to the collision cell by the gas pressure and gas type inside the collision cell. To control.

(3) Preferably, in (1) and (2) above , the sample to be analyzed is a mixture of a high molecular weight component and a low molecular weight component, and has a known molecular weight above the molecular weight of the high molecular weight component. A standard high molecular weight substance and a standard low molecular weight substance having a known molecular weight below the molecular weight of the low molecular weight component are ionized and introduced into the collision cell, and the control means has a maximum product ionic strength of the standard high molecular weight substance. The collision gas mixture ratio is stored, the collision gas mixture ratio at which the product ionic strength of the standard low molecular weight substance is maximum is stored, and the calibration curve is based on the two collision gas mixture ratios. Is created and stored, and the measurement sample is continuously measured while changing the mixture ratio of the collision gas along the calibration curve.

  ADVANTAGE OF THE INVENTION According to this invention, the mass spectrometer which can acquire the detailed structural information of a component to be analyzed can be implement | achieved, aiming at suppression of the enlargement of an apparatus and shortening of measurement time.

  In addition, according to the mass spectrometer of the present invention, each analysis target component can be cleaved under optimal MS / MS conditions, so that detailed structural information of the target component can be easily acquired.

  Further, MS / MS measurement from a low molecular weight component to a high molecular weight component can be performed by one analysis operation, and labor saving can be achieved.

  Embodiments of the present invention will be described below with reference to the accompanying drawings. In addition, embodiment of this invention is an example at the time of applying to an electrospray ion source as an ion source, and the mass spectrometer which uses 3DQ part for a collision cell.

  Further, in the embodiment of the present invention, an example will be described in which MS / MS analysis is performed on a mixed sample when the analyzed high molecular weight component is (A) and the low molecular weight component to be analyzed is (B). .

FIG. 1 is a schematic configuration diagram of a mass spectrometer according to an embodiment of the present invention.
In FIG. 1, an electrospray ion source 1 is ionized by applying a voltage and spraying on a sample to be analyzed, and introducing the ion into a 3DQ section. The 3DQ portion is formed by the end cap electrode 2, the ring electrode 3, and the insulator 4. The end cap electrode 2 and the ring electrode 3 are fixed and supported by an insulator 4.

  A gas introduction port 8 is formed in the insulator 4, and a gas introduction path 7 is connected to the gas introduction port 8. In the gas introduction path 7, a single pipe is connected to the gas introduction port 8, and the single pipe is branched into a plurality of pipes. Each of the branched pipes is connected to a collision gas generator 5 via a mass flow meter 6 for adjusting the gas flow rate into the 3DQ section.

  Each of the plurality of collision gas generators 5 is filled with different types of collision gases.

  The pressure in the 3DQ part is measured by a pressure sensor 9 arranged in the 3DQ part. The insulator 4 has an exhaust port 10 for adjusting the pressure in the 3DQ portion. A control valve 11 is disposed at the exhaust port 10, and the exhaust amount is adjusted by the degree of opening and closing of the control valve 11.

  The amount of preparation (open / closed degree) of the mass flow meter 6 is controlled by a control means (not shown). This control means is supplied with the pressure detection value from the pressure sensor 9, and controls the degree of opening and closing of the control valve 11 so as to become a predetermined pressure value according to the supplied pressure value.

  The ions in the 3DQ are discharged from the 3DQ to the detector 12 for each mass number, detected by the detector 12, and mass spectrum data is acquired.

  Ions derived from sample molecules introduced into the 3DQ portion from the ion source 1 are accumulated in the 3DQ for an arbitrary time set by the control means. At this time, by colliding the collision gas from the collision gas generator 5 so that the 3DQ has a constant pressure, a molecular cleavage reaction is caused by collision of the collision gas and molecular ions.

  Here, when only one kind of gas is used, the gas is introduced at a flow rate arbitrarily set only from the mass flow meter 6 of the used gas type, and the gas is not introduced from other lines.

  When a plurality of types of gases are mixed and used, the flow rate of each gas to be mixed is set by the mass flow meter 6 of the used gas, mixed in the path 7, and then introduced into 3DQ. Since the ion trap efficiency also changes when the type and mixing ratio of the collision gas change, it is necessary to optimize the amount of introduced gas each time.

  In the present invention, as described above, as a means for quickly setting the introduced gas amount to the optimum value, the flow rate of the mass flow meter 6 is adjusted, and the exhaust amount from the exhaust port 10 provided separately is also adjusted.

  Similar to the gas flow rate, the high-frequency voltage applied to the 3DQ electrode needs to be optimized depending on the collision gas type.

  In the present invention, after optimizing conditions for each sample using a plurality of standard samples with known molecular weights, a calibration curve is created, and the high-frequency voltage is changed over time in synchronization with the type of introduced gas. Set the optimum trapping conditions for the gas type.

  Of the separation columns used for sample separation, when a GPC column is used, the components in the sample are eluted in descending order of molecular weight. In the MS / MS method, a rare gas such as helium is generally used as a collision gas, but a high-mass rare gas such as xenon or argon having a large collision energy is used to cleave high molecular weight components. Thus, the reaction efficiency can be improved.

  However, since the collision energy is too large for low molecular weight components, the reaction efficiency decreases. However, the separation column of components such as GPC is used to separate the components, and the type of reaction gas used and the mixing depending on the elution time. By changing the ratio, each component can be efficiently cleaved even when the molecular weight distribution of the target component is wide. When a GPC column is used, the separation of the components in the sample depends on the molecular weight and elutes from the high mass components.

  In one embodiment of the present invention, the collision gas used for the reaction is initially set to xenon gas, and MS / MS measurement is started. Since the molecular weight of the component to be analyzed gradually decreases with time, helium is mixed with the gas to be introduced, and a gradient of the gas species ratio is created.

  According to this method, it is possible to efficiently cleave from a high molecular weight component to a low molecular weight component and obtain structural information of each molecule by one MS / MS measurement using a single collision cell.

  As another method, the same effect can be obtained by installing a gas inlet independently for each gas type and controlling the partial pressure of each gas type with the pressure in the collision cell being constant.

  Next, optimization of reaction gas species, optimization of gas flow rate, and optimization of trap voltage will be described by taking helium and xenon as collision gases.

1. FIG. 2 is an operation flowchart for optimizing the reaction gas species.
Ionization conditions are set in order to ensure the detection intensity of ions serving as precursor ions in the MS / MS analysis of each component.

  In step 201, a standard high molecular weight substance (X) and a standard low molecular weight substance (Y) whose molecular weight is predicted to be (A) having a molecular weight of (X) or less and (B) having a molecular weight of (Y) or more are transferred to a syringe pump. Then, the solution is continuously fed to the ion source 1 and ionized by the electrospray ionization method and introduced into the 3DQ portion.

  Next, in step 202, helium is allowed to flow into the 3DQ portion as a buffer gas for decelerating the collision gas and ions, but at this stage, the CID voltage for ion cleavage is not applied, and the precursor ions Detect intensity. The accumulation time of ions in 3DQ is arbitrarily adjusted, and the accumulation time at which the precursor ion intensity is maximized is confirmed and stored in the control means.

  Next, the operator monitors the product ions generated by MS / MS with the CID voltage applied to the 3DQ section turned on by the display means connected to the control means.

  The mass flow controllers 6 and 6 of the xenon gas channel and the helium gas channel are operated while confirming the spectrum pattern of the product ion of the high molecular weight substance (X) from the control means.

  The xenon / helium mixing ratio is changed while keeping the pressure in 3DQ constant. The gas ratio is stored in the control means with the condition that the target product ionic strength is maximized as the optimum condition, and the optimum condition is also stored for the low molecular weight substance (Y).

  A calibration curve is created based on these two gas ratios, and the gas change rate at the time of analysis is determined and stored in the control means.

  Next, in step 203, analysis of the actual sample to be analyzed in which the high molecular weight component (A) and the low molecular weight component (B) are mixed is started.

  From the start of measurement, the measurement is continuously performed while changing the gas ratio according to the mixing ratio of (X) and (Y) along the calibration curve set by the condition setting described above.

2. FIG. 3 is an operation flowchart for optimizing the gas flow rate.
As with the optimization of collision gas species, the optimization of the gas flow rate is performed for each of the standard high molecular weight substance (X) and the standard low molecular weight substance (Y) after confirming the precursor ion intensity.

  In step 301, the standard high molecular weight substance (X) and the standard low molecular weight substance (Y) are ionized and introduced into the 3DQ part.

  Next, in step 302, the reaction gas is introduced at the optimized gas mixing ratio. Then, in the state where the CID voltage is OFF, the mass chromatogram of the target ion m / z = x of the substance (X) is confirmed on the monitor screen, and the ion accumulation time that maximizes the ion intensity is set.

  Then, the CID voltage is turned ON, and a mass chromatogram of product ions m / z = x is confirmed.

  With the gas mixture ratio fixed, the setting of the mass flow meter (mass flow controller) 6 is changed to change the amount of introduced gas. The gas flow rate is stored in the control means (PC) with the condition that the target product ion intensity is maximized as the optimum condition, and is set as the analysis condition.

  Similarly, for the low molecular weight substance (Y), the optimum conditions for the gas flow rate are confirmed and stored.

  A calibration curve is created under the optimum conditions (X) and (Y) and stored in the control means.

  Next, in step 203, analysis of the actual sample to be analyzed in which the high molecular weight component (A) and the low molecular weight component (B) are mixed is started.

  From the start of measurement, the measurement is continuously performed while changing the gas flow rates of (X) and (Y) along the calibration curve set by the condition setting described above. In this case, the gas ratio is changed along the previously set calibration curve of the gas ratio.

3. Optimization of Trap Voltage FIG. 4 is an operation flowchart of trap voltage optimization.
Based on the previously optimized gas type and gas flow rate (impact gas introduction conditions), the trap conditions for each component are optimized.

  Step 401 is the same as steps 201 and 301.

  In step 402, no CID voltage is applied at the time of trapping. Therefore, gas is introduced under the gas introduction conditions for each component determined in advance, and the voltage value applied to the 3DQ section is scanned for (X) and (Y). The voltage value that maximizes the precursor ionic strength is determined as the optimum condition.

  Then, a calibration curve of voltage values is created under optimum conditions for each of (X) and (Y).

  The reaction conditions are controlled within the set analysis time by the calibration curves (shown in FIGS. 5A, 5B, and 5C) created by optimizing the gas species, gas flow rate, and trap voltage. Set on the means.

  Next, in step 403, analysis of the actual sample to be analyzed in which the high molecular weight component (A) and the low molecular weight component (B) are mixed is started.

  From the start of measurement, the measurement is continuously performed while changing the voltage value along the calibration curve set by the condition setting described above.

  The sample containing the high molecular weight component (A) and the low molecular weight component (B) is analyzed using the analysis conditions determined as described above.

  Next, in an embodiment of the present invention, a low flow rate (10 to 500 μL) liquid chromatograph and an electrospray ion source are used as sample separation and ionization means, and a GPC column is used as an example of a separation column. The results obtained are shown in FIG.

  The components in the mixed sample introduced into the separation column are separated by the separation column and sequentially eluted from the high molecular weight components. As shown in FIG. 6, at the start of analysis (T = 0), the collision gas is introduced at the optimum gas mixture ratio calculated based on the previously determined gas change rate, and the gas ratio is changed as the analysis proceeds ( In the example shown, the xenon / helium ratio is changed from 1 to 0 over time).

  By setting to the calculated mixing ratio until the end of analysis (T = t), MS / MS analysis can be performed continuously at the optimal collision gas mixing ratio for each molecular weight. It is possible to acquire structural information from a high molecular weight component to a low molecular weight component.

  In other words, when the mixing ratio is high, structural component information of a component having a high molecular weight is obtained, and when the mixing ratio is low, structural information of a component having a low molecular weight is continuously obtained in one analysis process (analysis time). Is obtained.

  In addition, when the slope of the calibration curve is steep or when it is necessary to suddenly increase or decrease the internal pressure of 3DQ every unit time, not only the introduction flow rate but also the gas flow rate discharged from the exhaust port 10 is controlled by the control valve. 11 can be accommodated by adjusting.

  For example, when it is necessary to rapidly reduce the internal pressure of 3DQ (assuming that the pressure is reduced from α to β), the actual pressure (α) detected from the pressure sensor 9 is confirmed by the control means, When the difference (α−β) from the required pressure (β) exceeds the set pressure difference (P), the control means can automatically open the control valve 11 to perform exhaust.

  In addition, as shown in FIG. 7, the present invention can use a quadrupole portion as a reaction field in a mass spectrometer in which a plurality of types of ion lenses such as the quadrupole mass analyzer 13 are connected. In the example shown in FIG. 7, the gas pipe 7 shown in FIG. 1 is applied to both the quadrupole part 13 and the 3DQ part 14, and a plurality of collision gas generators 5 are connected together with the mass flow meter 6. Further, the collision gas flow supplied to the quadrupole part 13 and the 3DQ part 14 can be cleaved under different conditions in the quadrupole part 13 and the 3DQ part 14 by individually controlling the mass flow meter 6. It is.

  In addition, by setting the standard sample molecular weight and necessary mass range to be used by the above control means, it is possible to easily calculate each calibration curve by introducing the standard sample and reflect it in the analysis conditions. It is.

  In the above-described example, two kinds of helium and xenon are used as the collision gas, but other gases can also be used.

  It is also possible to use helium, xenon, argon, and other three or more kinds of gases.

  In addition, according to the mass spectrometer of the present invention, detailed structural information can be obtained by a single measurement in protein analysis, synthetic polymer structure analysis, etc., in which the molecular weight distribution range of the target component is wide. Accuracy and labor saving are possible.

  Furthermore, the present invention can be applied to a gas chromatograph / mass spectrometer, a liquid chromatograph / mass spectrometer, etc. that are widely used for environmental analysis and the like, and can realize highly accurate analysis.

It is a schematic block diagram of the mass spectrometer which is one Embodiment of this invention. 5 is an operation flowchart for optimizing reaction gas species in an embodiment of the present invention. 5 is an operation flowchart for optimizing a gas flow rate in an embodiment of the present invention. 4 is an operation flowchart of trap voltage optimization in an embodiment of the present invention. FIG. 6 is a diagram showing a determined calibration curve in an embodiment of the present invention. In one Embodiment of this invention, it is a figure which shows the structure information obtained by changing the mixing ratio of mixed gas continuously with time. It is a figure which shows the example at the time of applying this invention to the mass spectrometer which connected the quadrupole mass spectrometry part and 3DQ part.

Explanation of symbols

DESCRIPTION OF SYMBOLS 1 Electrospray ion source 2 End cap electrode 3 Ring electrode 4 Insulator 5 Collision gas generator 6 Mass flow meter 7 Gas introduction path 8 Gas introduction port 9 Pressure sensor 10 Exhaust port 11 Control valve 12 Detector 13 Quadrupole mass spectrometry part 14 3DQ section

Claims (3)

In a mass spectrometer having a collision cell for performing a collision-induced cleavage reaction between a sample-derived ion and a collision gas, and mass-analyzing the ion,
A gas introduction path for introducing a plurality of types of collision gases into the collision cell alone or in combination;
A gas flow rate adjusting means for adjusting the flow rate of introduction of the plurality of types of collision gas into the collision cell for each of the plurality of types of collision gas;
Control means for controlling the gas flow rate adjusting means in order to adjust the time variation of the type, flow rate and collision gas mixing ratio of the collision gas introduced into the collision cell;
An exhaust port formed in the collision cell to adjust the pressure inside the collision cell;
A control valve for controlling the amount of exhaust gas from the exhaust port;
A mass spectrometer comprising: The mass spectrometer according to claim 1,
The mass spectrometer according to claim 1, wherein the control means controls an electric field applied to the collision cell and controls a voltage value applied to the collision cell by a gas pressure and a gas type inside the collision cell . The mass spectrometer according to any one of claims 1 and 2 ,
The sample to be analyzed is a mixture of high and low molecular weight components,
A standard high molecular weight substance having a known molecular weight above the molecular weight of the high molecular weight component and a standard low molecular weight substance having a known molecular weight below the molecular weight of the low molecular weight component are ionized and introduced into the collision cell,
The control means stores the mixture ratio of the collision gas that maximizes the product ion intensity of the standard high molecular weight substance, and stores the mixture ratio of the collision gas that maximizes the product ion intensity of the standard low molecular weight substance. A calibration curve is created and stored based on the mixture ratio of the collision gas at two points, and measurement is performed on the sample to be analyzed while changing the mixture ratio of the collision gas along the calibration curve. A mass spectrometer characterized by being performed continuously .

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