Mass Spectrometry Ionization Sources

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Mass Spectrometry Ionization Sources

Please check out our Mass Spectrometry section for more information or to find manufacturers that sell these products.

Whatever their configuration, mass spectrometers comprise two basic components: an ionization source and a mass analyzer. The former, as its name implies, ionizes molecules; the latter measures their mass—or more accurately, their mass-to-charge (m/z) ratio. Generally speaking, any mass analyzer can be coupled to any ionization source, and both are critical to a properly executed mass spec experiment. As a result, both must be considered when making a purchasing decision.

MALDI Ionization

Koichi Tanaka won a share of the 2002 Nobel Prize in chemistry for his discovery of this "soft" (that is, non-fragmenting) ionization technique. One of two such methods that opened mass spectrometry to biologists (the other being electrospray ionization), MALDI, or matrix-assisted, laser desorption ionization, is a solid-phase technique that uses laser energy to ionize molecules off a metal target plate.

Available from such vendors as Thermo Fisher Scientific, Life Technologies, and Shimadzu, among others, the process works like this: Sample is mixed with an organic matrix material (such as alpha-cyano-4-hydroxycinnamic acid), spotted onto the plate, and dried. The matrix is "tuned" to the laser; upon irradiation, it absorbs laser energy and heats up, essentially transferring its energy to the molecule of interest and producing ionized molecules capable of being detected.

The vast majority of ions produced in this manner have a charge of +1. That's because higher charged particles (such as +2 or +3), for the most part, "are energetically unstable," says Keith Waddell, LC-MS applications solution manager at  Agilent Technologies. He relates that MALDI pioneer and developer Michael Karas used to say, "In MALDI, the [singly charged] [M+H]+ ions are the lucky survivors."

According to Waddell, the technique favors polar compounds. "Generally, the more nitrogens in the molecule of interest, the more likely you are to get ionization. MALDI will also ionize hydroxyls and carbonyl compounds, but to lower efficiency. The more hydrophobic the molecule, the worse it will ionize in MALDI."

Electrospray Ionization (ESI)

John Fenn won his share of the 2002 chemistry Nobel for his discovery of another soft ionization method: electrospray ionization (ESI). Available on systems from Agilent, Shimadzu, Thermo, and Life Technologies, among others, ESI [like its sister techniques, atmospheric pressure chemical ionization (APCI) and atmospheric pressure photoionization (APPI)] is a liquid phase process that produces a fine mist of droplets, as from an atomizer or airbrusher. In all three cases, the sample inlet typically is coupled to an in-line liquid chromatography (LC) system; as a result, they are the mainstays of LC-MS and LC-MS/MS (though "electrospray is the single most popular ionization technique for LC-MS systems," according to Waddell).

What distinguishes the three techniques is the ionization process itself. In the case of ESI, as liquid exits the LC and passes into the sprayer, it becomes subjected to a "voltage offset" (between 2000 V and 3000 V) in the spray tip, says Waddell. This encourages a high electrical charge on the surface of the resulting droplets, which then pass into a heated environment full of dry nitrogen gas. As the droplets evaporate, "The ion density on the droplet surface eventually gets so high that charge transfers to the molecule, and the molecular ion gets lifted out," he says.

The process works in both positive and negative ion modes, and unlike MALDI, produces ions with a wide range of charge states (e.g., +1, +2, +3, …), making it amenable to high molecular weight species. Polar molecules (such as biopolymers) ionize especially well by ESI. "Nitrogen-containing compounds like amino acids, peptides, and oligonucleotides will ionize preferentially," says Waddell.

APCI Ionization

APCI, or atmospheric pressure chemical ionization, is an ESI variant that uses a "corona discharge"—basically, an electrified needle—to induce ionization of the solvent, which in turn reacts with the sample molecules to induce a chemical reaction resulting in an ionized sample molecule.

"In APCI, you rely wholly on a discharge across the spray to act on forming a chemical reaction, such that a charge is transferred to the molecule," says Agilent's Waddell (the company sells both standalone APCI and dual function ESI/APCI sources). "In other words, you cause chemical ions to interact with the molecule to cause the ionization process."

According to Waddell, APCI tends to favor relatively small, neutral, or hydrophobic compounds, such as steroids, lipids, and non-polar drugs, typically imparting a charge of +1.

APPI Ionization

Another ESI variant is APPI, or atmospheric pressure photoionization. According to Jonathan McNally, a marketing manager at Thermo Fisher Scientific (which offers a wide diversity of ionization sources, including a dual function APPI/APCI source), "APPI uses the same setup as APCI, but it doesn't use a coronal discharge to ionize the compounds; it uses photons." Specifically, high-intensity ultraviolet light ionizes the solvent gas, which in turn ionizes the sample molecules.

According to Waddell, one problem with APPI is that it is inefficient, and highly solvent-dependent. "You have to add toluene [or hexane] to the solvent that you are using on your LC in order to facilitate the APPI process," he says.

APPI ionizes liquid-phase compounds that are refractory to both ESI and APCI, such as highly non-polar molecules like napthols and anthracenes, says Waddell. But it is not for the environmentally conscious, he adds. "Because you are literally spraying toluene or hexane directly into the ion source, you need really good safety procedures to prevent poisoning the operators. It's thought to be unhealthy."

Electron Ionization (EI)

While ESI, APCI, and APPI are used strictly with LC, EI, or electron impact ionization, is a gas chromatographic (GC) ionization technique.

The technique, says Waddell, "is the granddaddy of them all." In use since the early 20th century, EI "relies on a glowing filament as in a light bulb," he says.

McNally explains: "The electrons bombard the molecules at about 70 eV, and that shoots out an electron from the compound, leaving a positively charged molecule. It usually fragments the compound as well, giving you a whole range of fragments, which you can interpret according to classic interpretation rules."

Best of all, the fragmentation patterns are unique and reproducible, such that researchers have been able to compile EI fragmentation reference libraries. Agilent, LECO, and Thermo all offer EI (and CI, below) sources on their GC/MS systems.

Chemical Ionization (CI)

CI, or chemical ionization, is also used exclusively for GC applications. Related to EI as APCI is to ESI, "Chemical ionization has an additional gas in the ion source," says McNally; typical gasses include methane, isobutane, or ammonia. The gas gets protonated by the EI filament, protonating sample ions in turn to +1.

According to Waddell, CI has the advantage of being somewhat less harsh than EI. "It was originally used to generate [parent] molecular ions for GC/MS," he says. That's because EI "generally fragments the molecule, and it's possible to not see a molecular ion." However, he adds, CI is also "less sensitive than EI," and presents additional maintenance and technical challenges.

"It's generally a more difficult technique than EI, but it does give you molecular ion information, and that, for a lot of people, is important."

Inductively Coupled Plasma Ionization

ICP, or inductively coupled plasma ionization, is by far the harshest ionization technique of the bunch. Says Waddell, "You are basically introducing a liquid and applying a blow torch to it." The idea, he explains, is to reduce a sample down to its atomic components.

"It's for elemental analysis," says McNally, and especially metals—for instance, to search for mercury in fish or lead in paint. "It gives the stoichiometry of a material," McNally says, "and can be used to determine the elemental composition of unknown compounds."

There are other ionization sources, of course—including, for instance: DESI (desorption electrospray ionization), an atmospheric-pressure, matrix-free variant of MALDI; DART (direct analysis in real time), which ionizes samples at atmospheric pressure using an electron beam; and LDTD (laser diode thermal desorption), which uses indirect laser energy to vaporize a sample that is then ionized via APCI. There are also numerous variants available, including so-called "nano-ESI," a popular proteomics source that applies ESI to extremely slow flow capillary LC.

As any given instrument typically contains only one (or maybe two) ionization sources, it's important to anticipate your applications before you buy. Says McNally, most sources "can ionize most things, the question is how efficiently it works. You always are looking for the most efficient way to ionize your compound."

Please check out our Mass Spectrometry section for more information or to find manufacturers that sell these products.