After ionization, analytes must be transferred from the source region at atmospheric pressure to a low pressure region of around 10-5 torr.48 This is achieved by a combination of a potential difference
down the ion path and pressure difference between the two regions.55 The ion plume in the
atmospheric pressure region is propelled by the electric potential difference towards the inlet of the low pressure region. As ions approach the inlet, the rapid acceleration of the bulk gas because of decreasing pressure causes a turbulent flow through the transfer capillary towards the differentially pumped regions. As the gas exits the ion transfer capillary, it rapidly expands into the low pressure region. A series of ion funnels, skimmers, and multipoles act to contain the ions as neutrals are pumped away. Throughout this process the ions are cooled, and off-axis velocity is dampened by electric fields until a beam of ions that is suitable for further analysis by a mass analyzer is
produced.56 The pressure reached is a function of the type of mass analysis performed – quadrupole-
based analysis operates at pressures around 10-4 torr whereas analyses requiring a long mean free
path (such as TOF or orbitrap experiments) operate at pressures up to 10-10 torr (a mean free path of
Figure 1.8. Resolution’s mass dependence on the Q-Exactive illustrated with three challenging to resolve A1 species. As mass increases instrument resolution decreases. This results in distinct mass peaks becoming more poorly resolved. (Green) 13C and 2H become indistinguishable at 550 Da. (Red) 13C and 15N become indistinguishable at 1000 Da. (Blue) 15N and 2H become indistinguishable at 1300 Da. Based on
experimentally observed resolutions at 280,000 resolving power.
1.5.1 The Q Exactive: Quadrupole-Orbitrap Mass Spectrometer
The Q Exactive (QE) mass spectrometer is a Fourier Transform (FT) based instrument that offers ultra-high mass resolving power, high mass accuracy and exquisite sensitivity. The mass spectrometer couples two mass analyzers, a quadrupole and an orbitrap, with a C-trap and collision cell intervening.57 The orbitrap is an ion-trapping device, into which a narrow beam of ions is
injected perpendicular too and off center from a central spindle electrode.58 Ions are confined
between the central electrode and the outer shell by electric fields, and begin to orbit perpendicular to the central electrode due to their initial velocity. The distribution of initial velocities and positions perpendicular to the central electrode cause the ion packet to spread into a ring around the central
electrode. Motion of the ion rings parallel to the central electrode follows naturally owing to the off- center injection of ions into the potential well. Ion motion along this parallel axis is dependent on the potential well and critically, the mass-to-charge ratio of the ions. (Equation 2) Thus, ion motion is observed as the image current in the two outer detector plates. This time domain signal can be transformed into a frequency domain spectrum (with frequency proportional to m/z) by FT.58
𝜔 = �𝑚/𝑧𝑘 (2)
Injection of ions into the orbitrap was a considerable design challenge – the initial distribution of position and momentum of ions parallel to the central electrode was limiting frequency
determination. Focusing of ions in this dimension is achieved by the C-trap, which upon injection compresses the ion packet into a narrow ribbon for injection. As the QE is a trapping instrument, observation of the ions occurs in a pulsed manner. Depending on the desired mass resolving power, orbitrap analysis can take as long as 1000 ms (for a resolving power of 256,000.)59 To minimize this
limitation’s effect on duty cycle, ions can be accumulated in the C-trap and fragmented in the collision cell with parallel acquisition of an orbitrap spectrum.
Even still, the orbitrap is inherently charge limited – as charge density in the orbitrap becomes too high dephasing of ion packets and saturation of signal amplifiers can occur. For this reason, duty cycle limitations for the QE are most often due to high ion flux rather than ion sampling limitations. This has an interesting practical result relevant to untargeted studies – the limit of detection is dependent on the ion flux during a particular scan. When a particularly abundant group of ions elute they occupy a large fraction of the charge capacity of the orbitrap. Thusly, the limit of detection for the entire scan is increased because fewer of other species will be accumulated.
The charge limitations of the orbitrap mass analyzer necessitate a rationing of space in the trap. Rationing is accomplished by quadrupole mass filtering prior to ion accumulation. In the low
abundance case, when ion flux is limited, the orbitrap is an exquisitely sensitive instrument, with comparable sensitivity to triple-quadrupole type instruments. Similar to the triple quadrupole, the QE offers nearly 100% duty cycle when not charge saturated. Additionally, the QE is able to observe ions for an extended amount of time and offers high mass resolving power of possible interferences in the monitored fragments. These factors allow the QE to equal and exceed the triple quadrupole in targeted sensitivity.
Figure 1.9. A schematic of the Q-Exactive mass spectrometer.*
1.5.2 The Quadrupole-Time-of-Flight Mass Spectrometer
The Quadrupole-Time of Flight Mass Spectrometer (QTOF) is a hybrid instrument coupling a quadrupole and collision cell to a time-of-flight mass analyzer.60 Time-of-flight mass analysis is
achieved by a high voltage pulse accelerating a packet of ions into a drift region. This pulse imparts the same amount of kinetic energy to each ion, but ions with different mass-to-charge ratios will travel at different speeds owing to the conservation of momentum. As such, ions are separated
based on the time it takes them to travel a several meter distance and detected upon impacting a detector plate. (Equation 3) Differences in the initial positions and velocities of the ions contribute to peak broadening, this is mitigated by the use of a repulsive electrostatic region that reflects the ions back in the direction they came from, focusing each mass packet.61
𝑡 = 𝑘�𝑚/𝑧 (3)
The QTOF mass spectrometer is a pulsed instrument, requiring drifting ions to reach the detector prior to the next pulse of ions. As such, duty cycle on these instruments is around 10%. Improvements to this include using Hammond transforms and overlapped pulses, as well as gating of the ions. As opposed to the QE, the QTOF has a very large charge capacity, and is limited primarily by detector saturation. As such, other ions in the spectrum have no impact on the overall sensitivity of a scan and the QTOF is suitable for bright ion sources.
Figure 1.10. A schematic of quadrupole-time-of-flight mass spectrometer.*