INAUGURAL ARTICLE by a Recently Elected Academy Member:Gas-phase ions of solute species from charged droplets of solutions

For the past several decades, the production of gas-phase solute ions by solvent evaporation from charged droplets of solution, as in electrospray ionization (ESI), has enjoyed increasingly widespread use in the detection and identification of the complex organic molecules of interest and importance in biochemical systems (1). Even so, there is still much debate on the mechanism(s) by which those gaseous ions are formed. The two most favored of these mechanisms are embodied in the charged residue model (CRM) originally proposed by Malcolm Dole et al. in 1968 (2) and 1970 (3) and the ion evaporation model (IEM) suggested by Iribarne and Thomson in 1975 (4). The CRM has its roots in a theoretical paper published by Lord Rayleigh (John Williams Strutts) in 1882 (5). In that paper, Rayleigh addressed the question of what would happen as solvent evaporates from a droplet of volatile liquid containing an excess of either anions or cations. He reasoned that the repulsive forces between those excess charges of like sign would cause their associated ions to be situated at equidistant intervals on the surface of the droplet. As the droplet size decreased by evaporation of solvent, those surface ions would get closer and closer together until the integral over the droplet surface of the coulomb repulsion forces between those surface ions would exceed the integral of surface tension of the droplet liquid over that same area. At that point, now frequently referred to as the “Rayleigh limit,” the droplet would increase the available surface area by breaking up into a plurality of smaller offspring droplets. These offspring droplets also would undergo solvent evaporation until they, too, would reach the Rayleigh limit and subdivide into still smaller droplets. Such subdivision of evaporating charged droplets had indeed been observed and reported by Zeleny (6). Dole et al.'s (2, 3) idea was that a succession of such subdivisions of the original droplets would eventually lead to the formation of “ultimate droplets” so small that each of them would contain only one solute molecule. As the last solvent molecules evaporated from such an ultimate droplet, the residual solute molecule would retain some or all of the charges on that droplet to become a gas-phase solute ion. They also realized that this scenario might make possible the production of gas-phase ions of molecules too large to be ionized by the then-customary procedures based on gas-phase encounters between neutral molecules and sufficiently energetic electrons, photons, or other ions. The large oligomer molecules in which they were interested simply could not be vaporized by the usual methods without undergoing catastrophic thermal decomposition. In 1968 and 1970, Dole et al. (2, 3) published the results of their attempts to produce and mass analyze ions of polystyrene oligomers by means of this ESI technique. For several reasons, not realized until later, the molecular weight values they obtained were at odds with what was known about the probable values for those oligomers. Consequently, their results did not persuade other investigators to follow their lead until some years later. Fortunately, as it turned out, Dole himself did live long enough to see those ideas evolve into what is now sometimes referred to as “The Electrospray Revolution” by which ESI-MS has become one of the most widely practiced analytical techniques now in use. Sometime after Dole et al.'s first two papers (2, 3) on ESI, Iribarne and Thomson (4) had experimented briefly with the technique and in 1975 offered a somewhat different explanation for the possible production of gas-phase ions by evaporation of solvent from charged liquid droplets (4). They argued that before a charged droplet became small enough to contain only one solute molecule, the charge density on its surface would become so high that the resulting field would be sufficiently intense to “push” one or more of those surface ions into the ambient gas, thereby forming gaseous ions of at least some of those solute molecules. Continued evaporation of solvent from successive generations of such charged droplets thus would ultimately result in driving many, if not most, of the surface cations (or anions) on the original droplets into the gas phase. The diagrams in Fig. 1 attempt to illustrate schematically these two possible mechanisms by which nonvolatile solute species in a charged droplet could become free ions in the ambient gas. Fig. 1. A schematic representation of the possible pathways for ion formation from a charged liquid droplet. The upper and the lower parts of the diagram illustrate the ion formation mechanisms depicted in the CRM of Dole et al. (2, 3) and the IEM of Iribarne ... In the years after Dole et al.'s papers (2, 3), there were several attempts to develop a satisfactory ionization technique based on these charged droplet scenarios but none of them were very successful. Then in the 1980s, building on the ideas of Dole et al. (2, 3) and Iribarne and Thomson (4), and drawing on our own extensive studies on the free jet expansion of gases from relatively high pressure into vacuum, our group, then at Yale, found the right combination of conditions and demonstrated the successful production and mass analysis of intact solute ions from molecules having a wide range of molecular weights (7, 8). This “success” has spawned much argument and discussion on the mechanisms by which such ions could be formed (9–21). Over the past few years, we have investigated this ion formation mechanism for a number of molecular species having a wide range of compositions and molecular weights, e.g., tetra-alkyl ammonium compounds, amino acids, peptides, proteins, carbohydrates, and some polar synthetic polymers such as polyethylene glycols (PEGs) (12, 13). After careful consideration of all of the results, we conclude that for most, if not all, cases in which ESI is effective, gas-phase solute ions are formed from charged droplets according to the sequence of events described in the IEM of Iribarne and Thomson (4). However, in the case of very large parent species, including PEGs with molecular masses as high as 5,000,000 Da, we believe that the CRM of Dole et al. (2, 3) may comprise the more likely ionization scenario (13). Even so, recent studies by de la Mora (16) suggest that ions of globular proteins (nondenatured) with molecular masses as low as 6,600 Da may also be produced according to the CRM. Indeed, some species seem capable of forming multiply charged ions by either the CRM or the IEM but de la Mora and Gamero-Castrano (17) argue persuasively that smaller species like tetraheptyl ammonium cations are usually formed by the IEM. By a somewhat different approach, Kebarle et al. (18, 19) have also extensively investigated the mechanisms of ion formation for a variety of solute species including metal atoms and protein molecules. They concluded that metal ions such as Na+, K+, or Cs+ are most likely to be produced by the IEM (18), whereas larger ions of denatured proteins probably are produced by the CRM (19). More recently, Williams and colleagues (20, 21) have studied the formation of highly protonated ions of peptides, proteins, and synthetic polymers when the electrosprayed solutions were doped with compounds having low vapor pressures, e.g., glycerol or m-nitrobenzyl alcohol. The results from such studies led them to conclude that organic ions, with a mass range from 100 to 7,000 Da, were most likely produced by the CRM (21). In this article, we present experimental results that seem to provide positive proof that the actual mechanisms by which electrospray ions are most often produced from charged droplets is embodied in the IEM of Iribarne and Thomson (4). We hope these results may serve to resolve some of the remaining uncertainties about those mechanisms.