MALDI can very efficiently generate the gas phase ions from a variety of non-volatile and thermolabile molecules such as proteins, carbohydrates, nucleic acids, and synthetic polymers. MALDI can desorb and ionize the molecules as large as 300 kDa. MALDI usually results in the molecular species having only one charge. In positive ion mode, a quasimolecular ion is formed by protonation of the molecule (MH⁺). The samples or the matrices can have trace amounts of alkali metal ions, often resulting in the quasimolecular species, MNa⁺ or sometimes MK⁺ (Figure 11.8). Multiply charged species are also observed sometimes.

 

Figure 11.8 MALDI-TOF mass spectrum of a peptide with theoretical mass of 789.464 Da. The peaks at 812.4442 and 828.4162 represent [peptide–Na+] and [peptide–K+], respectively.

Time-of-flight (TOF) mass analyzers are well suited for the pulsed ionization techniques such as MALDI (In MALDI, laser pulses produce the ions in pulses). We shall be discussing the different mass analyzers in the next lecture.

Electrospray ionization (ESI)

In electrospray ionization, the analyte solution enters the ionization chamber, maintained at atmospheric pressure, through a fine capillary. The typical flow rates are ~1-20 μl/min. A potential difference of ~3-6 kV is applied between the capillary and the counter-electrode that is ~0.3 – 2 cm away. Under this electric field, the sample droplets appearing at the capillary end accumulate large amount of charge. If the potential of the capillary is above a threshold voltage, the drop will be dispersed into a very fine spray. A coaxial sheath is present around the capillary through which dry nitrogen is supplied for better nebulization and restricting the dispersion of the spray in space. Evaporation of the solvent causes the droplets to diminish in size and their charge density to increase (Figure 11.9). The high charge density on these droplets can further result in the production of smaller droplets. The droplets keep losing the solvent ultimately resulting in the desorption of molecular ions from the surface. As the ions are generated from the surface, a surface active molecule will be detected better. For large molecules such as proteins, the molecules do not desorb from the droplets but become ionized through complete evaporation of the solvent.

Figure 11.9 Diagrammatic representation of ionization in an ESI ion source

 

ESI typically results in multiply-charged molecular species. Biological macromolecules usually result in the mass spectra wherein the consecutive peaks differ by one charge unit. This information allows one to calculate the mass of the biomolecule.