Catherine J. McNeal
Focus
Department of Chemistry Texas A&M University College Station, Tex. 77843
obtained when there is a near-grazing collision of the ion or atom beam with the solid or liquid surface. The sput tering yield reaches a maximum near angles of 15° to 20° for impinging ions having energies on the order of 1-20 keV. Below 15° the particles begin to penetrate only into shallow layers, and much energy is reflected via high-en ergy particles which are not detected, making energy dissipation less effi cient. Donald Sedgwick, a chemist at Manchester Institute of Science and Technology, England, and codeveloper of FABMS, corroborated Sigmund's statements, reporting that in their laboratory molecular ion yields did in crease as θ approached more oblique angles and that the optimal angle was found to be 15°. Sedgwick also added that at "head-on" incident angles, more fragmentation was observed in the spectra. Similar results were also reported by Bo Sundqvist, a physicist at the Tandem Accelerator, Uppsala, Sweden, using fast (ca. 50 MeV) heavy ions from an accelerator, but he added that no qualitative differences were observed in the heavy ion induced de sorption spectra as θ approached 90°. One of the primary differences de scribed by Sigmund, Ken Standing, a physicist from the University of Mani toba, Canada, and Sundqvist in their presentations was that if the imping ing particle has an energy below ca.
Symposium, on East Atom and Ion Induced Mass Spectrometry of Nonvolatile Organic Solids A unique ensemble of scientists, ranging from physicist to biologist, gathered at the Johns Hopkins School of Medicine on Sept. 11 for a sympo sium on fast atom and ion-induced mass spectrometry of nonvolatile or ganic solids. The meeting was spon sored by the Middle Atlantic Mass Spectrometry (MAMS) Laboratory with support from Kratos Scientific Instruments and VG Instruments, Inc. Six invited speakers from Europe, Canada, and the U.S. discussed the mechanisms of molecular and frag ment ion production and the various parameters affecting ion yields. The list of speakers was almost uniformly divided between those using low-ener gy (keV) primary ion (or atom) beams [e.g., fast atom bombardment mass spectrometry (FABMS) and secon dary ion mass spectrometry (SIMS)] and those using high-energy (MeV) particles [e.g., heavy ion induced mass spectrometry (HIIDMS) and 262Cfplasma desorption mass spectrometry (252Cf-PDMS)]. Throughout the pre sentations the similarities and differ ences in secondary ion yields (both molecular and fragment ions) and ion ization and desorption mechanisms between these two approaches were described. Both the theoretical foun dations and the experimental observa tions were addressed. During the round-table discussion the following day the six speakers and a dozen invit ed scientists, primarily from mass spectrometry laboratories on the East
0003-2700/81 /0351-043A$01.00/0 © 1981 American Chemical Society
Coast, discussed the effects of the in cident beam angle, the desorption mechanism, and the fragmentation mechanisms of large molecules. Peter Sigmund, a physicist from Odense University, Odense, Denmark, led the round-table discussion on the effects of the angle of incidence of the incoming particle beam. This angle, Θ, is defined to be the angle between the surface and the primary ion beam. For a beam parallel to the sample surface θ = 0°. Sigmund stated that the sput tering yield (yield of desorbed ions and molecules) varies with cos Θ. Max imum yields should, theoretically, be
500 -g· 400 c · : 3oo ο
(dEldx),
% 200 (dE/dx)n |
1 MeV/amu
1 keV/amu
100 0 0"4
10~3
10"2
10~1
10°
101
102
103
Ε (MeV)
Figure 1. Energy deposition density (dE/dx) for 35CI impinging on 27 AI. The (dE/dx) curve for nuclear excitation is designated by {dE/dx)n. The electronic excitation curve is designated as (dE/dx)e. From P. K. Haff, "Erosion of Surfaces by Fast Heavy Ions" in "Heavy Ion Science," Vol. III., D. A. Bromley, Ed., Plenum, in press, with permission. Courtesy of K. Standing ANALYTICAL CHEMISTRY, VOL. 54, NO. 1, JANUARY 1982 · 43 A
Focus
252
Cf PDMS
HMD
100 Me V Tc
SIMS
20 keV Cs +
Solid Film
FAB
2 KeV Ar°
Liquid Drop
LD
Laser Pulse
Figure 2. Schematic diagram of HIIDMS, SIMS, FABMS, and LDMS. From R. Macfarlane, Ace. Chem. Res., in press 44 A · ANALYTICAL CHEMISTRY, VOL. 54, NO. 1, JANUARY 1982
0.1 MeV/amu most of its energy loss in the sample occurs through elastic collisions with the atomic nuclei of the sample molecules, whereas in the MeV-range energy is lost by electronic excitation. Both Standing and Sundqvist showed graphical comparisons of these two regions. The rate at which a charged particle loses energy in pass ing through matter (dE/dx) is plotted as a function of beam energy in Figure 1. These theoretical curves were plot ted for chlorine ions impinging on an aluminum target. The scaling factors should be similar for heavy ion (> Ζ = 6) or heavy atom bombardment of or ganic films. Sundqvist and Standing both reported that the yield of molec ular ion species increased with in creasing projectile ion energy and with increasingly larger projectiles. In the keV energy range secondary ion yields are enhanced as the mass of the pro jectile increases. In the MeV range the secondary ion yields scale as the square of the atomic number. Further differences in the effects of MeV vs. keV projectile energies on the ionization and desorption mechanisms were discussed by Sigmund, Sundqvist, and Ronald Macfarlane, a physical chemist at Texas A&M Uni versity. Each described how for low velocity (keV) particles the energy is dissipated in the sample substrate by generation of a collisional cascade. With MeV projectiles the energy is dissipated primarily via electronic ex citation and ultimately through lattice vibrations. Both mechanisms ulti mately give rise to a thermally activat ed region with the core attaining tem peratures near ΙΟ4 Κ. Macfarlane pointed out that the reason enhanced yields are observed in going to more massive projectiles is because the radi al dimensions of the core increase. The temperature remains constant. He also stated that desorption and ioniza tion mechanisms of laser desorption mass spectrometry (LDMS) are com patible with this thermal description and suggested that the generation of a hot spot is the central feature of or ganic SIMS, FABMS, LDMS, and HIIDMS. This global picture is indi cated pictorially in Figure 2. Direct comparisons of the same sample irradiated with keV and MeV projectiles were presented by Sundqvist. He reported the data of Duck, Treu, Voit, Albers, and Wein, who studied valine positive molecular and fragment ion yields as a function of the 1 6 0 projectile energy. These re sults are shown in Figure 3. The two maxima correspond to 10 keV and
Focus
(M-COOH)+ 6
I
(M + H) + 2
0
0.1
0.2 0.5 1.0 Velocity (cm/ns)
2.0
Figure 3. Valine (M + H) + and (M—COOH) + yields as a function of the 16 0 beam velocity. From P. Duck, W. Treu, H. Voit, A. Albers, and K. Wien; Proceedings from the Nordic Symposium on Ion Desorption of Molecules from Bioorganic Solids. B. Sundqvist, Ed., "Nuclear Instruments and Methods," in press, with permission. Courtesy of B. Sundqvist
8 MeV and are very close to the two maxima in the energy deposition density function (Figure 1). Sundqvist also noted the data of his colleagues Kamensky and Hâkansson who compared the molecular ion yields for a series of compounds of increasing molecular weight up to ca. 2000 u for 3 keV Cs+ ions and 54 MeV Cu+ 4 ions. (The symbol u, formerly amu, is defined as 1/12 the mass of an atom of C-12.) These results are presented in Table I. As in the Duck study, molecular ion yields were enhanced by utilizing projectiles having energies in the MeV range. This effect is apparently even more pronounced as the mass of the
sample increases. They suggested that there may be a threshold level for energy deposition in the organic film if molecular ions of relatively large species are to be produced. One additional difference between MeV and keV mass spectrometry is that for the former, Sundqvist reported that there was a strong charge-state dependence. He showed that for glycylglycine (M + H ) + ions the yield varied closely as q 4 where q is the charge on the projectile ion. Sigmund stated that for low-energy particles, no charge-state dependence is expected. The most striking difference between FABMS and the other methods discussed was the use of a liquid phase (e.g., glycerol) containing the sample molecules instead of the thin, solid film employed in the other techniques. As Sedgwick pointed out, the advantage to this is that the molecular surface is continuously refreshed, thereby avoiding surface damage that in a solid film would lead to quenching of the ion yields. One of the unique results presented in this vein was the effect of the solution concentration. Sedgwick pointed out that for neomycin sulfate, constant molecular ion yields were obtained until evaporation of the glycerol was complete, at which point production of sample ions ceased. The mass spectrum he obtained was therefore completely independent of the compound concentration. However, for the neutral neomycin no molecular ions were initially observed. It was only after a part of the glycerol evaporated that yields increased and finally reached a steady maximum in the time remaining before the solvent completely evaporated. Sedgwick suggested that in the case of the neomycin sulfate the sample was on the surface the entire time, whereas for the free base the compound appeared on the surface only after the concentration increased. In this case abundant glycerol ions are initially observed, verifying that only the surface molecules become ionized
Table I. Ratio of Yields for High-Energy Vs. Low-Energy Bombardment3
a
Compound
Isotopically averaged mass (u)
Glycylglycine (M + H)+ Ergosterol (M+) Bleomycin-B1 + Cu (Mt) MMTrAl'tTCEiA^TCEJbzCl;
132 397 1375 1884
(Courtesy I. Kamensky, P. Hâkansson, and B. Sundqvist)
(MeV/keV) Yield
13.7 ± 18.2 ± 200 ± 196 ±
.5 .5 25 50
and desorbed. Frank Field, a physical chemist from Rockefeller University, suggested that these data reflect that a chemical effect rather than a concentration effect is involved. He proposed that a protonated acidic species that could protonate neomycin could be formed from glycerol during the bombardment. In general there were many similarities in the mass spectra produced by the ion- and atom-induced desorption techniques. Macfarlane confirmed that the HIIDMS spectra were identical to the 252 Cf-PDMS spectra in all cases. Standing and Field verified that the organic SIMS and 252 Cf-PDMS spectra were also similar. Although no spectra were shown at the symposium of a common compound run by SIMS, FABMS, and 252 Cf-PDMS, representative results from each technique are shown in Figures 4, 5, and 6. Sedgwick showed the FABMS spectra of several peptides, the largest of which was of the β chain of insulin (M = 3495 u). A useful feature of the peptide spectra is that the amino acid sequence can be deduced from the fragmentation pattern. Figure 4 shows a mass-analyzed ion kinetic energy spectrum (MIKES) that records all ions formed from a protonated angio tensin (M + H ) + precursor. Sedgwick used this spectrum to demonstrate that the positive fragment ions origi nate from the (M + H ) + species and therefore are the result of gas-phase fragmentations and are not pyrolysis products. He pointed out that the fragmentation mechanism involves protonation of the nitrogen atom in one of the peptide bonds followed by rupture of that peptide bond. Occur rence of this process at multiple sites gives rise to the pattern in Figure 4. Only singly charged ions were ob served in the angiotensin spectrum. Doubly charged ions were said to be rarely observed; however, some had been detected in a spectrum of bradykinin. Sedgwick also reported that the sodium content of the sample greatly influences the amount of fragmenta tion. If sodium is added to the sample at a level such that the cationized species (M + Na) + is preferred over (M + H ) + production, fragment ions to a large extent disappear. In the round-table discussion Field reported that in his 252 Cf-PDMS studies of peptides, the (M + H)+ peak in the time-of-flight spectrum is substantial ly broader than the (M + Na) + peak. When deflector plates are employed in the measurement to filter out neutrals or metastable products, the (M + H ) + peak width is reduced to close to that of the (M + Na) + peak. Field con-
ANALYTICAL CHEMISTRY, VOL. 54, NO. 1, JANUARY 1982 · 47 A
Focus 1182+ π = 3
η = 4
η = 5
η = 6
η = 8
η = 9
1296+
+
+
+
1296+
1296 +
1296
\
1296
506 +
Ι
Ι
Ι
!
343+
1296
756 +
619 +
1000+
η = 7
η = 10 1296 +
•
1250+
Ι
1137+
ι
1296+
Ι
853+
Figure 4. MIKE spectrum of protonated angiotensin 1 (H-Asp-Arg-Val-Tyr-lle-His-Pro-Phe-His-Leu-OH) produced by FABMS. η = 1 for Asp, η = 2 for Arg, etc. Courtesy of D. Sedgwick
Mol Wt 2079.0
(M + Nay z i υζ [M + Η)*
2080 2022
1135
1x8
683
(M - TCE)1947
1361
(M - H)2078 1621 647
944 -Cl Figure 5. Secondary ion mass spectra of a chemically blocked ribotrinucleoside di phosphate. MMT = monomethoxytrityl, Bz = benzoyl, Sil = fert-butyldimethylsilyl, C Bz = benzoylcytosine, A Bz = benzoyladenine, U = uracil. Courtesy of K. Standing 48 A · ANALYTICAL CHEMISTRY, VOL. 54, NO. 1, JANUARY 1982
curred that, as in FABMS, it is the (M + H ) + species that tends to frag ment and not the "natriated" species. He also added that more fragmenta tion is observed when lithium rather than sodium is present in the sample matrix. Peptide spectra presented by Standing were comparable to the FAB spectra. The molecular ion was ob served in good intensity, and a similar fragmentation pattern was obtained. Standing also showed the SIM spec trum of a fully protected ribooligonucleotide (M = 2079 u), which is the largest species to be analyzed thus far by his laboratory (Figure 5). Both the positive and negative ion spectra were remarkably similar to the HIID and the 252 Cf-PD mass spectra. Negative fragment ions, from which the base se quence could be determined, were clearly apparent. The fragmentation spectrum is less complex than that of the peptides, thereby permitting facile and unambiguous identification of the primary structure. Figure 6 is the positive ion 252 Cf-PD spectrum of a fully protected deoxyoligonucleotide containing 14 residues. The cationized molecular ion is appar ent at mlz 6980. A second ion at mlz 5261 was determined to be an impuri ty. Macfarlane reported that this is the largest monomeric species ob served in the laboratory at Texas A&M. Sundqvist showed several examples of high molecular weight compounds analyzed in his laboratory. He re ported that peptides having molecular weights in excess of 6000 u have been detected as well as the dimeric species, demonstrating that the HIID method can be utilized in the mass range well beyond mlz 10 000. One of the points made early in the
Focus
6980
2250 2000 £· 1750
5261
1 1500 α> ΐ 1250 1000 750 500 250 5000
6000
7000
8000
9000
Mass {mlz) Figure 6. 252 Cf-plasma desorption positive ion spectrum of a chemically blocked deoxyoligonucleotide. The chemical structure is indicated above the spectrum. Vertical lines represent the deoxyribose moiety. DmTr = dimethoxytrityl, R = p-chlorophenyl, Iso = isobutyrl, bz = benzoyl. Courtesy of C. McNeal, R. Macfarlane
symposium by Robert Cotter of the Johns Hopkins School of Medicine was that spectra can be obtained by thermal desorption of simple com pounds that were thought to be "invo latile." As an example he showed the spectrum of glucose containing NaCI (Figure 7) that was obtained by pass ing a current (1-4 A) through the probe tip. The observed ions are all natriated species. No electron beam was used to effect ionization nor was a high extracting field used at the
source. He stated that at temperatures as low as 500-700 °C, ions of quater nary amines could be desorbed. More energy was required for producing the cationized molecular ions than for desorbing the quaternary ions, but this was less than that required for the production of molecular ions by pro tonation or by electron abstraction. This order of "ease of ionization" (R 4 N+, (M + Na)+, (H + H)+, Mf ) is significantly similar to that observed in other desorption methods (LDMS,
IN8+
100ι L 80
(M - 3H 2 0 + Na) + 149
I-
(M + Na) + 203
20 109 125
K+ 0 20
40
60
80
100
120
167 140
160
185 180
200
mie Figure 7. Thermal desorption mass spectrum of glucose containing NaCI. From R. Cotter, Anal. Chem. 1981, 53, 1307 50 A · ANALYTICAL CHEMISTRY, VOL. 54, NO. 1, JANUARY 1982
252
Cf-PDMS, FABMS, SIMS, FDMS) as to suggest a thermal basis to these methods where the Langmuir-Saha prediction of a distribution of desorbed ions and neutrals reflects the fact that lattice energies of ionic species are typically lower than ioniza tion potentials. Macfarlane suggested that, based on Cotter's results, the definition of a thermally labile molecule should be changed. He emphasized that if intact molecules or ions can be vaporized rather than the decomposition prod ucts, then the species is not thermally labile. He suggested that for molecular ions to be observed, the lifetime of the heated area (or hot spot) must be less than the time at which vibrational ex citation of the bond (leading to de composition) occurs. He believed this was a common feature of all the meth ods described. Additional topics discussed were the fragmentation patterns for large molecules, mechanisms of surface ion ization, and the recent observation at the U.S. Naval Research Laboratories of cesium iodide clusters extending beyond mlz 18 000. There was much lively discussion between theorist and experimentalist and between "lowenergy" and "high-energy" scientists. Participants agreed that the sympo sium provided an excellent forum for the exchange of information from di verse disciplines that helped to pro vide a more coherent basis for atom and ion induced mass spectrometry.
