community because Thermospray had become the technique of choice. The complexities of the interface and the limitations that the technique imposed on the LC solvent composition however, meant that the technique remained predominantly in the hand of the mass spectrometry community.


However, a number of minor events and developments were happening in tandem across the world which, when brought together, would change the LC-MS landscape completely. It is not often in science, that events coincide in this way, but this is an example one of those wonderful periods. So let me try to catalogue the events. Firstly, we find that Jack Henion (of DLI fame circa 1978) who was still working in the veterinary medicine arena at Cornell University, linked up with Bruce Thomson (of ion evaporation fame circa 1978) who was then working for a small mass spectrometry company called Sciex in Toronto who built air monitoring mass spectrometers. They published a seminal paper in Analytical Chemistry titled ‘Determination of sulfa drugs in biological fluids by LC-MSMS’ [14] which used atmospheric pressure chemical ionisation in front of the Sciex mass spectrometer. This approach to LC-MS demonstrated incredible sensitivity at trace levels. This was quickly followed by improvements in the design of the APCI source and further papers by Henion et al [15,16] soon after. Around the same time John Fenn at Yale University, had developed the first atmospheric pressure electrospray source based on the original work of Dole back in the 1960s [17]; and he reported on the multicharging of large bioorganic molecules such as proteins (see Figure 5) [18].


Figure 6. LC-MS analysis of three sulphonamides; MS spectrum shows the predominant MH+ from peak 2 at 4.49 minutes.


Figure 5. API Electrospray MS spectrum of the protein Cytochrom C.


So suddenly and by two independent research groups the world had been exposed to two techniques both using ionisation at atmospheric pressure; both showing incredible sensitivity, and both capable of linking to reverse phase LC. It was also fortunate that through Thomson the triangle was able to be completed, because he had access to an instrument company with a mass spectrometer which sampled at atmospheric pressure, so we had what could be called a marriage made in heaven.


The first publications started appearing in the literature under the titles of ‘ion spray’ and ‘APCI’ [19] and all were demonstrating sensitivities at least one to two orders of magnitude greater than other techniques. This was particularly interesting to the drug metabolism industry and by the time Sciex launched their first instrument in 1989, the whole landscape of LC-MS changed irreversibly. This new breed of instruments were simple to use, had no real constraints on mobile phase composition or flow rate; the spectra were simple to interprete, as the protonated molecular ion predominated the spectrum (Figure 6), with virtually no fragmentation being detected; even proteins and their digests could be analysed.


All of this meant that LC-MS was no longer wholly the domain of the mass spectrometrist and so biochemists, drug metabolism specialists and chromatographers, could all use these systems. In a very small space of time all the other mass spectrometer manufacturers follow suit and produced instruments exclusively dedicated to these API techniques. Without the amazing development of Atmospheric Pressure Ionisation we would not be where we are today. For example API led to the development of ‘open access’ LC-MS within the pharmaceutical industry. Mass directed purification was also a result of this development.


This incredible sensitivity means that API LC-MS is almost exclusively used in the drug metabolism community to quantify metabolites at low levels. The technique has also revolutionised the detection of trace level contaminants in drug formulations. Outside the pharmaceutical industry, the technique is centre stage in the detection of trace levels of performance enhancing drug within the sports community, and in the horse racing arena. Without it the life science community would not have progressed in the many ‘omic fields. The protein elucidation field would not have progressed to where it is today, for example the elucidation of large non covalently bound protein complexes is critically dependant on this approach. The list of API application areas is endless. It would not be an exaggeration to say that through the development of atmospheric pressure ionisation techniques, LC-MS has become a central mainstay within the analytical community. In my opinion it is probably the key tool in the analytical chemists toolbox, and one that we now take for granted.


Back in 1974, Patrick Arpino drew this now famous cartoon (Figure 7), which showed how unlikely it was that two apparently incompatible techniques could end up happily married together, but, within 30 years it was achieved.


I will leave you with one thought before I finish. The next time that you run an LC-MS instrument, or sit down with others to review some LC-MS data, just spend a short time and reflect on all the groundbreaking work and all the heartache that went into the development of this technique by a few pioneers who had the dogged resolve to overcome what was seen at the time to be an insolvable problem.


Analytical Science is a fascinating discipline to be a part of, and this story is one that should be recognised as a real achievement of success against enormous odds.


References


1. V.L. Tal’roze, G.V. Karpov Russian J. Phys. Chem. 42 1968, 1658-1664.


2. M.A Baldwin, F.W. McLafferty Org. Mass Spectrom., 7 1973, 1111-1112.


3. A Melera, Proceedings of 1979 Pittsburgh Conference paper number 85.


4. J.D. Henion Anal. Chem., 50 (12) 1978, 1687-1693.


5. P.J. Arpino, G. Guiochon, P. Krien, G. Devant J. Chromatogr., 185 1979, 529-547.


6. D.I. Carroll, I. Dzidic, R.N. Stillwell, K.D. Haegele, E. Horning, Anal. Chem. 47 (14) 1975, 2369-2373.


7. W.H. McFadden, H.L. Schwartz, S. Evans, J. Chromatogr. 122 1976, 389- 396.


8. D.S.Millington, D.York Adv. Mass Spectrom. 8 1980, 1819-1822.


9. R.D. Smith, A.A. Johnson Anal. Chem. 53 (7) 1981, 1120-1122.


10. B.A. Thomson, J.V. Iribarne J. Chem. Phys. 64, 1976, 2287-2294 .


11. C.R. Blakley, M.J. McAdams, M.L. Vestal, J. Chromatogr. 158 1978, 261- 276.


12. R.C. Willoughby, R.F. Browner Anal. Chem. 56 (14) 1984, 2626-2631.


13. Y. Ito, T. Takeuchi, D. Ishii, M. Goto J. Chromatogr. A. 346 1985, 161-166.


14. J.D. Henion, B.A. Thomson, P.H. Dawson Anal. Chem. 54 (3), 1982, 451-456.


15. T.R. Covey, E.D. Lee, J.D. Henion Anal. Chem. 58 (12) 1986, 2453-2460.


16. L.O. Weidolf, E.D. Lee, J.D. Henion Biomed. Environ. Mass Spectrom. 15 1987, 283-289.


17. M. Dole, L. Ferguson, M. Alice J. Chem. Phys. 49 1968, 2240-2249.


Figure 7. LC and MS, will the two ever come together? Highlighting the incompatibility between liquid and gas phase (vacuum based) systems - Patrick Arpino 1974.


18. M. Yamashita, J.B. Fenn J. Phys. Chem. 88 (20) 1984, 4671–4675.


19. A.P. Bruins, T.R. Covey, J.D. Henion Anal. Chem. 59 (22) 1987, 2642-2646.


Chromatography Focus


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