傅里叶质谱教程（英文）.docx

Part I - FTMS Tutorial
Motion of a Charged Particle in a Uniform Magnetic Field:
The basic principles of cyclotron motion are based on the fact that the ions of interest are situated in a uniform magnetic field. An ion’s flight path may or may not be deflected depending on which direction the particle is traveling in relation to the direction of the magnetic field.

Case 1 demonstrates a situation where an ion’s velocity, shown by the vector in red, and the magnetic field, shown by the vector in green, are moving parallel to each other in the z-direction. In this case the direction of the ion is not affected in any way by the magnetic field.

In Case 2 the ion, shown again in red, is moving perpendicular to the direction of the magnetic field. When this situation occurs there is a force that is exerted on the ion which causes it to be deflected into circular orbits. This force is given by the following equation:

F = q (v × B)
F = force on the ion
q = charge on the ion
v = velocity of the ion
B = magnetic field

Notice that in the expression we are taking the cross product of the velocity and the magnetic field. When calculating a cross product a factor of sin (where is the angle between the velocity vector and the magnetic field vector) is introduced. Since the velocity and magnetic field vectors are perpendicular in this case, the final cross product will be perpendicular to v and B, and its magnitude the simply the product q v B. This force is shown by the vector in blue. A positively charged particle will be deflected into an orbit in the clockwise direction, while a negatively charged particle will be deflected into a counterclockwise direction.
Cyclotron Motion:

In the diagram above we are looking down the magnetic field in the bore of our horizontal magnet. Inside this bore is a charged particle moving perpendicular to the magnetic field, which is directed straight off the page at you. The two components which make up this force are the magnetic force, the inward force on the ion (F1), and the centrifuge force, the outward force on the ion (F2). In order to keep the ion in a stable circular orbit the magnetic force and the centrifuge force must exactly equal each other.

By equating the two forces we can derive that the angular frequency () of an ion with mass m and charge q:

This angular frequency is called the cyclotron frequency in these applications. It is important to note that in this final equation the cyclotron frequency has no dependence of the velocity of the ion. Thus ions of a given mass will have the same cyclotron frequency, regardless of the time the ion enters the cell or the velocity with which the ion enters the cell. This allows us to avoid the peak broadening seen with time-of-flight mass spectrometers and achieve ultra-high mass resolution.
Example Calculations:

As an example of the mathematics, we will go through a simple calculation.

Usually, cyclotron frequencies are expressed in Hertz (cycles per second) rather than radians per second. Dividing the cyclotron frequency by 2 radians/cycle gives the frequency, f, in units of Hertz.

f = /2 = 6.7542 x 10^5/2
f = 107.45 kHz

Thus the singly charged ion is spinning inside the analyzer cell at about 100,000 times per second. . Note that if the ion had contained 4 charges to begin with, we would need to multiply our final answer by 4 to get the correct value. So, in this case the cyclotron frequency would be about 428 kHz, or 428,000 cycles per second if there had been in the +4 charge state.
There is a much easier formula to remember that already has all of the conversion factors figured in:

Note: This formula does assume that the ion is singly charged.
Image Current Detection:
FTMS is unique in its method of detecting ions and deriving a mass spectrum. This method is based on the ion cyclotron resonance (ICR) principle. We will take ions produced by electron ionization, electrospray ionization or matrix-assisted laser-desorption ionization and store these ions inside an ICR analyzer cell. This cell is situated in the homogenous field region of a large superconducting magnet, and the ions will be moving (as discussed previously) in circular orbits at their cyclotron frequencies. Most mass spectrometers use an electron multiplier or photomultiplier to detect a signal, but FTMS uses a resonance method (that is completely analogous to NMR) to detect the image current signal generated by the ions in the cell.

Above is a diagram of an ion cycling in the magnetic field between two electrode plates. Normally the radius of an ion’s orbit will be about 0.1 mm, but if we send in an RF frequency to the cell that is equal to the cyclotron frequency of the ion, it will gain energy from the RF field and spin up to a larger orbit. The plates in slide 6 can be viewed as the receiver plates in our analyzer cell. As a positively charged ion circles near to electrode 1, it will induce electrons to collect on the electrode. Then, as the ion circles over near to electrode 2, the electrons migrate back to collect on the second electrode instead. This back-and-forth migration of the electrons is converted to an image current by placing a resistor on the wire connecting the two electrodes, and the resulting image current is a sinusoidal signal. The signal at this point has a size on the order of microamps, so we also run the signal through an amplifier to raise its size to the order of milliamps, and then we send this output to a computer. FTMS can typically detect thousands of ions in the analyzer cell simultaneously.
Fourier Signal Analysis:

If you had multiple ions of the different mass in the cell, they would all be spinning with similar orbital radii but different frequencies. Thus, the overall signal coming out of the cell is actually a composite signal (a superposition) of all of the cyclotron frequencies of all of the ions present. A good analogy for this is to think of the individual m/z ratios of the ions we are looking at as the keys of a piano. Any one key has a frequency that corresponds to the cyclotron frequency of one of our ions. If you hit just one key at a time, our brains can pick out exactly which key is being pressed. On the other hand, if you took a large hammer and suddenly smashed the keyboard of the piano, this act will set all of the strings vibrating, as though we had hit all of the keys at once. Then we get a loud burst of sound, but our brains will not be able to pick out exactly which of the keys were “hit” at a given time. However, using a computer and the mathematics of Fourier transform methods, we can deconvolute the overall signal to display exactly which frequencies were contributing to the total signal.

In terms of our mass spectrometer, what we do first is “slam shut the door to the keyboard” by accelerating all the ions with an RF sweep (commonly called a chirp) that covers a range of frequencies corresponding to the m/z values we wish to observe. This will accelerate all the ions in the cell near to the plates of the cell, and then we can wait and measure the image current transient that results from this excitation. This transient signal will be a composite of the cyclotron frequencies of all the ions present in the cell, and the signal slowly dies off with time as the ions relax and return their stable circular orbits in the center of the analyzer cell. This signal is digitized and stored in our computer, and a Fast Fourier Transform (FFT) algorithm is used to extract the frequency and amplitude for each component of the composite signal. This frequency plot is then converted to an m/z plot and displayed as the mass spectrum.
Part II - FTMS Tutorial
Advantages of the IonSpec Fourier Transform mass spectrometers:
Now that we know a little bit about how the ions are moving inside our analyzer cell, and some of the basic concepts about FTMS detection, the question arises of why we would want to use FTMS? Three main reasons are:
o Mass resolution in excess of 800,000
o Attomole sensitivity
o Less than 1 ppm mass accuracy
These are direct benefits of using Fourier Transform methods to derive the mass spectra. We should note that the high resolution and mass accuracy we can achieve are a direct consequence of the fact that our cyclotron frequencies have no dependence on the velocity of the ions in the analyzer cell or the radius of their orbit. It is also important to remember that resolution and mass accuracy both increase linearly with the strength of the magnetic field used. Our electrospray unit is the most sensitive of any mass spectrometer available, and our ability to reach ppm mass accuracy is critical when trying to identify an unknown.
One of the key features of the Explorer system is the quick switch that can be done between the MALDI/EI and Electrospray vacuum carts. It is simply a matter of rolling one vacuum cart out of the superconducting magnet and rolling the other vacuum cart into the magnet from the other side. The switchover can be done in less than 2 minutes. Both carts take advantage of our patented RF quadrupole ion guide which efficiently transports ions from the source region, through the magnetic field, to the analyzer cell. The fastest commercially available computers are used so that we can calculate large Fast Fourier Transforms (FFT’s), and our windows-based software is very easy to learn and use. Even novice users can learn and use the software in less than a day. Over the last 19 years the electronics have become continuously more modern and compact, with fewer cables, fewer connections, and more integration for increased reliability. Installation is accomplished in less than a week.
Ease of Use:

The above picture is of IonSpec’s main line product, the Explorer FT Mass Spectrometer. This instrument leads the way as the highest performance mass spectrometer available. However, the Explorer FTMS is actually two mass spectrometers in one. Positioned on the right side of the superconducting magnet is the Electrospray vacuum system, and positioned on the left side of the superconducting magnet is the MALDI vacuum system. The two vacuum systems are totally separate and independent from each other. The MALDI vacuum system has its own set of pumps and its own set of electronics, and the same is true for the Electrospray. However, both vacuum systems are controlled by the Omega Data Station. Switching between the MALDI and Electrospray ionization sources is extremely fast and efficient because the two vacuum carts are mounted on wheels, so they are able to roll in and out of the magnet very easily. Having two separate vacuum carts is highly efficient for all types of laboratory environments. In a service laboratory it is impossible to know whether ESI or MALDI samples will be sent in for analysis. With the Explorer FTMS, the instrument operator can be running ESI samples while the MALDI samples are being prepped. When it comes time to switch the operator simply rolls the ESI vacuum system out of the magnet and rolls the MALDI vacuum system in. The rest of the switching is taken care of by the OMEGA Data Station.
EI and MALDI Source:
Now we are going to move on and start talking in more detail about each of the carts. The first cart that we will discuss is the combined electron impact and MALDI (matrix-assisted laser desorption ionization) source cart. EI and MALDI are combined in the same cart because in both cases the ions are generated in high vacuum.

For MALDI, we have a laser that will be beamed in with mirrors and lenses to a sample plate. Ions will be blasted off of the sample plate, into the source region that is pumped down to about 10-6 Torr. Those ions are then electrostatically pulled through a mechanical shutter and into the RF quadrupole ion guide. The shutter used by IonSpec maintains a lower pressure in the analyzer cell which results in high mass resolution. The ions pass down the center of the quadrupole ion guide and are injected into the center of the FTMS cell. The ions are trapped in the analyzer cell by pulsing the voltages on the trapping plate. This method puts the ions at the center of the magnetic field, in the most homogeneous region, which results in higher mass resolution than the "kick sideways" method used by another manufacture. It is the fringing fields of the magnet that require us to use a quadrupole ion guide. Without the ion guide, the fringing fields would repel the ions, and they would be deflected away from the analyzer cell.
Various stages of pumping are present. The two pumps nearest the analyzer cell can be either cryogenic pumps or turbomolecular pumps. In the source region we would use a split-turbomolecular pump. All of the turbomolecular pumps used would be backed by one mechanical pump. This system features revolving powers in excess of 800,000, mass accuracy on the order of 1 ppm and sensitivity on the order of 1 femtomole. We can do MS/MS on the ions that are generated to get structural information out of those ions. We also have data base searching tools available for people who are doing protein and peptide work. A variety of magnets are available: 4.7T, 7.0T or 9.4T, and 12.0T actively-shielded magnets.
Ion Injection:

Ion injection is a key step, so we will discuss it in more detail. A schematic of the RF quadrupole ion guide is shown above. The quadrupole ion guide is able to inject a broad mass range, and it collimates the ion beam tightly in the x,y direction so that the ions are kept at the center of the quadrupole. As a result, the ions are at the center of the cell after injection and ready to be manipulated as desired. It is not necessary to pre-cool the ions down to the center of the cell. The RF ion guide has the highest ion transfer efficiency and lowest injection energy (<10 eV). All the major FTMS research labs and ThermoFinnigan use the same RF ion guide method as IonSpec. Another nice feature of the quadrupole guide is that it can be run with very low voltages, so we are not accelerating the ions in the z direction to an appreciable extent. The ion guide is usually set to anywhere between 5 and 15 volts. This very gentle voltage is able to get the ions to move all the way down to the cell while the quadrupole field keeps the ions trapped in their trajectories and pushes them through the magnetic field without being deflected. The RF quadrupole ion guide is quite easy to use. Other than the single adjustable voltage, it has two operating frequencies that allow us to run at a lower (100-1000 m/z) and a higher (400-2500 m/z) mass range.
Since we hold the patent on this technology, other instrument vendors are not able to use the quadrupole ion guide. As a result, they have to smash the ions through the magnetic field quickly to keep them from being deflected away from their analyzer cell. This requires high voltages of a couple thou