The force exerted on ions placed in an area exposed to constant and uniform magnetic fields moves those ions in a circular orbit that is perpendicular to the direction of the magnetic field.98 An ion-cyclotron resonance (ICR) analyzer is a cubic cell (~2 cm each edge), consisting of two opposite trapping plates, two opposite excitation plates, and two opposite receiver plates. The cell is placed between the poles of an electromagnet or superconducting solenoid magnet with typical field strengths in the 3 to 7 T range (Figure 2.12). Ions, which may be formed by almost any currently used ion source, become trapped upon introduction into the cell and begin to "cyclotron," or move in circular orbits.
The frequency of the circular motion (natural cyclotron frequency, fc) is directly proportional to the strength of the magnetic field (B) and inversely proportional to the m/z ratio of the ions, according to the equation vc = 2nfc = v/r = kzB/m, where fc is frequency, v is ion velocity, r is the radius of the orbiting circle, and k is a proportionality constant. When an rf field (a frequency-swept "chirp" signal) is superimposed perpendicular to the direction of the magnetic field, those ions with cyclotron frequencies equal to the excitation frequency will absorb energy from the rf field and move into orbits of larger radii. These ions are translationally excited and move coherently, i.e., in phase with the exciting field, between the receiver plates. When an external conducting network is attached to the receiving plates, the ions transmit a complex rf signal that contains frequency components related to their m/z values. When a group of coherently moving positive ions approaches one of the receiver plates the ions attract electrons thus creating a current; as they continue moving on their orbit and approach the other plate, they again attract electrons. (Negative ions move in the opposite direction and repel electrons as they approach the receiving plates.) The image current signal begins to decay as the coherency is disturbed over time. The complex timedomain image currents thus produced can be transformed into frequency-domain signals by Fourier transform analysis (FT) to yield the component frequencies of the different ions from which mass spectra can be obtained using the equation given above.98
The operation of FTICR is governed by a series of computer-controlled discrete events, known as experimental pulse sequence. There are three categories of individual events. First: application of d.c. or rf excitation to increase the amplitude of the cyclotron motion, in order to: (a) establish coherent high-amplitude ion packets for detection; (b) eject ions from the cell; (c) facilitate CID.
Second: altering trap plate potentials, in order to: (a) inject or eject ions along magnetic field lines or (b) manipulate the z-amplitude of the ions. Third: delaying events, that occur by not making changes in the electric fields and allowing time for: (a) relaxation (homogeneous and inhomoge-nous); (b) collisions (dissociative and reactive); and (c) detection of the image current.
There are four steps common to all FTICR experiments: (1) establish a large electric field gradient between the trap plates; during this quench period all trapped charged particles are removed from the cell; (2) inject ions from an external ion source or ionize sample within the cell; (3) excite the ions; and (4) detect ions by image current. Mass scanning can be accomplished by varying the rf pulses (frequency of irradiation) at a fixed magnetic field.98,99
There is a trend toward using higher magnetic fields, the main advantage of which is improved detection of high-mass ions. This is because of the increased cyclotron frequency associated with these magnets, which makes the slower moving high mass ions easier to detect by avoiding the environmental noise found in the low frequency region.100,101 Other advantages include improvements in resolution and detection efficiency102 and routine mass measurements to part-per-million accuracy.103
Because of the low drift velocity of the ions and the long cycloidal paths, the actual time it takes for the ions to traverse the analyzer is 5 to 10 ms in contrast to the few |js flight times in other analyzer types. The low drift velocity necessitates operation in very low vacuum, typically in the 10-10 torr region. The long flight times of ions mean that a gas pulse (the type of gas may be varied) that raises the pressure to 10-6 torr is typical for CID experiments. This amount of gas is readily pumped out of the cell, making these instruments particularly suited for MSn experiments (Section 2.5).
FTICRMS has several unique advantages: (a) extremely high mass resolution, up to 3 million, may be achieved because m/z values are calculated from cyclotron frequency determinations that can be made to nine significant figures; (b) several masses may be detected simultaneously; (c) MS/MS experiments up to MS4 can be conducted; and (d) change from one operational mode to another, e.g., from high to low resolution or from full mass spectra to multiple ion detection, can be made by changing only electronic parameters. Limitations include a drop in resolution with increasing mass (although the mass range is up to 10 kDa or higher), moderate dynamic range, and difficulties in quantification. Despite the relatively simple mechanical structure of the ICR cells, the need for high intensity magnetic fields, ultrahigh vacuum, and sophisticated computer techniques make the instrumentation expensive.
FTICR is perhaps the most promising of current MS techniques. Advantages include high-accuracy (±0.001%) molecular mass determinations of biopolymers when in combination with ESI or MALDI.100,104 Improved ion transmission efficiency from ESI sources with an electrodynamic ion funnel, together with some other technological advances,105 resulted in LOD of ~30 zmol (~18,000 molecules) for proteins in the 8 to 20 kDa range.106 The field has been reviewed extensively both with respect to principles99,107 and applications.108-110
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