Ionization of Large Molecules

Mass spectrometers measure the mass-to-charge ratio (m/z) of ions. They consist of an ionization source that converts molecules into gas-phase ions and a mass analyzer coupled to an ion detector to determine the m/z ratio of the ion (Yates III, 2000). A mass analyzer uses a physical property such as time-of-flight (TOF) to separate ions of a particular m/z value that then strike the detector (Fig. 2.3). The magnitude of the current that is produced at the detector as a function of time is used to determine the m/z value of the ion. While mass spectrometers have been used for many years for chemistry applications, it was the development of reproducible techniques to create ions of large molecules that made the method appropriate for proteomics. 

Matrix Assisted Laser Desorption Ionization. During the development of MS, a lot of studies have been devoted to the use of laser light as an energy source for ionizing molecules. As a result, in the mid 1980s MALDI[5] was introduced and soon applied to the study of large molecules.[18] Koichi Tanaka was jointly awarded the Nobel Prize for Chemistry in 2002 for the study of large biomolecules by MALDI. 

In 1988-1989, two processes were discovered that allowed the transfer of large molecules into the gas phase matrix-assisted laser desorption and ionization (MALDI) and electrospray ionization (Karas and Hillenkamp, 1988 Fenn et al., 1989). 

Since then, it has proven to be one of the most successful ionization methods for the mass spectrometric investigation of large molecules, increasing the... 

Ionization is very important in that you have to have ions to detect and measure mass. Today, electrospray ionization is used this is a process where a streaming liquid is ionized and this imparts ionization of the molecules dissolved within it. However, large amounts of liquid or uncharged molecules are not desirable in a mass spectrometer operating in a vacuum. Hence, electrospray ionization is optimized such that nearly all of the solvent/spray is removed and only charged molecules enter the mass-separation devices. You may ask how I ensure that only the molecules I am interested in make it to the mass spectrometer rather than all of the other molecules that may be present in the mix. This is an important concept, since... 

The experiments demonstrate that femtosecond laser pulses offer new opportunities for multiple-photon ionization of bioorganic molecules on surface. The fast femtosecond excitation makes it possible to produce molecular and fragmentation ions directly on the surface being irradiated. The two-photon excitation with an intense femtosecond pulse allows the selectivity of ionization of the chromophore (tryptophan in our case) in large molecules... 

Femtosecond photoionization mass spectrometry might be useful in the study of the three-dimensional structure of large biomolecules. When a selectively excitable and ionizable chromophore is located on the outer (surface) part of large molecule, one can be detached in the picosecond time scale. However, when the excitable chromophore is located in the inner part of the big molecule, its detachment will require a much longer time, which is needed for spatial rearrangement of the molecule. So, even the simple mass spectrometry of bioorganic molecules with femtosecond laser ionization can reveal some details of their spatial structure. 

A problem with all mass spectroscopy of large molecules is how to get them into the vapor phase so that they can be ionized and their fragmentation patterns determined. Simple heating may cause excessive degradation and formation of ions not corresponding to the desired substance. Two useful methods that involve only intense short-term local heating of the sample appear to have promise in this connection. One method uses a burst from a powerful infrared laser to volatilize part of the sample, and the other uses bombardment by heavy and energetic particles from fission of californium-252 nuclei to raise the local temperature of the sample to about 10,000°. The latter technique both volatilizes and ionizes the sample molecules. 

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