The renaissance of time-of-flight mass spectrometry

international Journal of Mass Spectrometry and Zon Processes, 99 (1990) l-39

Elsevier Science Publishers B.V., Amsterdam

THE RENAISSANCE OF TIME-OF-FLIGHT MASS SPECTROMETRY*

D. PRICE and G.J. MILNES Department of Chemistry and Applied Chemistry, University of Salford, Salford M5 4 WT (U.K.)

(Received 6 October 1989)

ABSTRACT Although the time-of-flight mass spectrometer based on the Wiley-McLaren pulsed twogrid ion source has been available since the early 1960s its application has been limited by low resolving power and sensitivity. The renaissance of interest in this instrument is, in part, a consequence of the development of new ionisation techniques such as plasma and laser desorption, which are advantaged by the time-of-flight spectrometer’s unique ability to provide a complete mass spectrum per event, and its high mass range. This has led to design developments which give major improvements in instrument performance which promise to yield rich rewards particularly in the study of large biomolecules.

INTRODUCTION

This article is based on review lectures given at the dynamic mass spectrometry (DMS) symposia held in 1986 at Canterbury and recently Salford. It essentially covers the literature since our last review was published in 1984 [l]. In the intervening period a number of accounts of time-of-flight (TOF) mass spectrometry have appeared. Most importantly, Campana has edited a special issue of Analytical Instrumentation devoted to TOF mass spectrometry [2]. As well as containing an historical overview and an account of design theory, this issue contains articles on linear, reflectron and hybrid instruments, together with a report of a TOF mass spectrometry workshop concerned with current and future developments. This volume is an excellent starting point for anyone considering entering the TOF mass spectrometry arena. A review of TOF mass spectrometry with particular reference to its use with desorption probes is provided by Le Beyec [3] whilst Brunnee [4] discusses its contribution within an overall view of mass spectrometry. In contrast with some of our earlier TOF mass spectrometry articles, the problem with * Paperpresented at the 10th Triennial International U.K., 3-5 July 1989. 0168-l 176/90/$03.50

Mass Spectrometry Symposium, Salford,

0 1990 Elsevier Science Publishers B.V.

2 TABLE 1 Advantages of the time-of-flight mass spectrometer Complete mass spectrum for each ionisation event-spectra can be obtained for very small amounts of sample (approaching attomole) Ideal where ionisation is pulsed or spatially confined High transmission Unlimited mass range Fastest scanning mass spectrometer; repetition rates up to 100 kHz Performance dependent on electronics rather than mechanical alignment Relatively low cost

this article has been what to omit from the vast number of TOF mass spectrometry papers now appearing in the literature. Inevitably the choices made show a personal bias. To those who feel their work ought to have been included we apologise; to those who read this article with a view to entering the TOF mass spectrometry field we say welcome to a technique which promises rich rewards to come. The current decade has seen a renaissance in the development and utilisation of the TOF mass spectrometry technique, with many new and different types of instruments becoming availabPe. As a result, TOF mass spectrometry is now beginning to make a really significant impact in the world of mass spectrometry. This is because its unique advantages of being capable of providing a complete spectrum per event combined with unlimited mass range make it the best choice of mass analyser for the newer ionisation techniques such as plasma and laser desorption currently being developed and which promise rich rewards in the study of large biomolecules. The advantages of the time-of-flight mass spectrometer (TOFMS) are summarised in Table 1. RESOLUTION

PROBLEM AND FOCUSING

TECHNIQUES

The limited mass resolution of the conventional Wiley-McLaren [5] type of TOFMS was the principal reason why it never established more than a small share of the mass spectrometer market. Considerable effort is currently being expended to overcome this limitation. The approaches taken improve the resolving power either by modifying the operation of the Wiley-McLaren instrument or by conceiving new instrument design. Opsal et al. [6] have reviewed the factors which limit mass resolution and have developed a model, based on probability theory, for the calculation of ion peak shapes and mass resolution. Model calculations indicated that mass resolutions in the TOFMS can be greatly increased by reducing the velocity spread of the ions. This was confirmed experimentally when, using supersonic molecular beam cooling of the sample prior to ionisation, a resolution of 4650 (FWHM) was obtained for benzene. Good agreement was obtained between the experimental data ob-

3

tained under space focusing conditions and the calculated peak profiles. The most common method of improving the resolution of the TOFMS is the application of so-called reflectron techniques (decelerating and reflecting fields) originated by Mamyrin et al. [7]. For ions of the same m/z ratio entering these fields, those of higher energy penetrate further before reflection than do slower lower energy ions. Consequently, the faster ions have a longer flight time through the reflection than do the slower ions. This effect compensates for the reverse behaviour across the linear flight paths of the instrument. The appropriate reflection voltages can be adjusted so that the fast and slow ions reach the detector together, thus optimising resolution. The original reflectron of Mamyrin et al. [7] was designed with bent ion-beam geometry. This led to difticulties with focusing the ion beam in the angles of emission from the source and required an analyser chamber of fairly large diameter. To circumvent these disadvantages, Mamyrin and Shmikk [8] have since designed a linear reflectron which enables the improvement in resolution to be obtained with a small instrument. Shmikk [9] has described the performance of an instrument based on this design. Practical details of a linear mass reflectron based on the design of Mamyrin and Schmikk have been published by Lubman et al. [lo]. In the conventional design of Mamyrin and Shmikk [8] the decelerating and reflecting fields are separated by various grids. Use of a gridless reflectron should result in increased ion transmission and thus enhance sensitivity. There are several theoretical approaches to such a design [I 1,121, and Frey et al. have recently designed and constructed such an instrument [13]. Glashchenko et al. [14] have suggested a technique for calculating the axial potential distribution of an electrostatic mirror for which the total ion drift time is independent of the ion energy. This would make it possible to specify an electric field strength to ensure TOF focusing of ions of specified energy. Instruments based on the original design of Mamyrin et al. [7] have achieved resolving powers of 10000 or better. Detailed consideration of the field distortions in the vicinity of wire meshes indicates that the original restrictions on the lengths of the retarding and reflecting fields suggested by Mamyrin for the two-stage reflectron limit the resolution obtainable to around 10 000 [ 151.Calculations show that much higher resolutions should be accessible by increasing the retarding Geld length compared with that suggested by Mamyrin. This approach has been taken by Bergmann et al. [16] who have recently given a preliminary report on the development of a time-offlight mass analyser (TOFMA) with a mass resolution of 35 000 for Cs atoms. A retarding field segment 29cm long is used in the two-stage reflectron. The latter has been designed to have good rigidity and very low temperature sensitivity in order to obtain very stable and homogeneous electric fields. More detailed descriptions of the ion optics and reflector design have been promised in future publications.

4 R/N6 CLECTRODES

PARASOUb wz,=$z

NE10

,z

DE TEnOR

Fig. 1. Sketch of a TOFMS in which the whole of the flight path occurs within a parabolic reflection field controlled via voltages on the ring electrodes. (From ref. 4, Fig. 71.)

A modified reflectron mass spectrometer using UV-laser-induced desorption ionisation via prism internal reflection has been reported by Yang and Reilly [ 171.Theoretical calculations indicate that mass resolution greater than 1000 000 should be possible if a picosecond laser and appropriate electronics are available. Experimentally, a value of 11000 at m/z 93 was achieved using a nanosecond laser. With a view to reducing the overall size of the instrument, Su [18] has developed a TOFMS with two sets of parallel-plate electrostatic fields to enable multiple reflection of ions. A prototype instrument with 20cm lieldfree space had a resolving power of 3570 for 85Rb. Instrument designs are now appearing in which the whole of the ion flight path occurs within the reflectron [19,20] (Fig. 1). Theoretically, perfect time focusing should be obtainable in a parabolic field which can be generated via suitable control of the voltages at the ring electrodes. Whilst major improvement has undoubtedly followed the Mamyrin’s development of the reflectron, there is still a need to improve the performance of the conventional Wiley-McLaren type instruments in view of the large number in use. To this end Sanzone and co-workers [21] have described an impulse field focusing technique which should enhance the performance of TOFMSs of any conventional design. This involves the application of a time-dependent ion draw-out field as opposed to the time-independent field employed in conventional TOFMSs. Experiments have shown that the technique can result in considerable improvement in the resolution obtainable. Details of the circuitry used to generate the lOOV, nanosecond rise-time pulses required are also given [22].

Enke and co-workers [23J have considered various models for improved resolving power in TOFMS, A model for dynamic post-source acceleration was developed in an attempt to achieve mass-independent space and energy focusing. The model incorporated all pertinent parameters including the extraction voltage, the accelerating and field-free region lengths, and the dynamic acceleration function. Mathematical analysis and computer simulations based on equations from this model were used to develop a new postsource acceleration technique. Simulations indicated that concurrent space and energy focusing was possible, and that unit resolving power can be achieved up to at least 2000 Da using typical gaseous source conditions and readily attainable instrumental parameters. Kinsel and Johnston [24] have suggested post-source pulse focusing as a simple method to improve the resolution of a linear TOFMS. The gaining of a factor of at least 3 in resolution over a static field TOFMS should easily be achievable while still maintaining an inst~ental con~guration which can be used for a static field TOFMS. The method involves application of a focusing voltage pulse to a short field-free region located after the source and accelerating region. The pulse is timed so that it occurs after the ions of interest have entered this region. Instrumental requirements are identical with those for time-lag focusing but without the disadvantages of the latter. This technique should be particularly valuable for surface desorption/ionisation and heated sample introduction. In addition, various instrumental alterations are discussed which suggest possibilities for achieving resolving powers in excess of 10 000 for a linear TOFMS. One interesting approach to improving the performance of the WileyMcLaren type of TOFMS is the “velocity compaction” technique developed by Muga [25]_The principle of “velocity compaction” can be briefly described in the following manner. In each cycle of the instrument, ions are accelerated out of the ion source into a short deft-re~on where partial separation into different m/z packets occurs. These packets then enter a time-dependent and monotonically time-varying accelerating field region. The slower ions in each packet will be subjected to a higher voltage and hence greater acceleration than the faster ions. As a consequence, nearly all the ions in a packet will attain the same final drift velocity before entering the final drift-region in front of the detector. In this way the spatial distribution of ions within each m/z packet is “frozen” whilst peak-to-peak adjacent mass ion packets continue to separate during the final drift period with minimal overlap. This is in contrast with the situation in the conventional constant-voltage-acceleration TOFMS for which increasing overlap occurs during the final drift period. Simulation of a TOFMS, based on the “velocity compaction” principle and utilising varied accelerating voltages and optimised grid positions, indicates that resolving powers beyond 10 000 should be attainable. Modi~cation of an old Bendix Model 12 TOFMS to demonstrate the technique resulted in a

resolution of 1176 at 173 Da. A commercial TOFMS based on the velocity compaction principle is now available [26]. This has a mass range up to 2000Da with a resolution better than 2000 (FWHM). Increasing chromatographic resolution demands higher scan speeds for mass spectrometers used as chromatographic detectors. The potential of the TOFMS has not as yet been realised because of poor resolution. Also TOFMSs are not fully compatible with certain alternative modes of ionisation, e.g. chemical ionisation (CI) which cannot be pulsed sufficiently rapidly for normal operation [27]. Pinkston et al. [28] have reported work aimed at solving this problem by introducing an electrostatic energy analyser between the ion source and a linear TOFMA. Beam deflection plates are located just before the flight tube so that ion admission can be pulsed. The advantages of this arrangement are the improved resolution resulting from the introduction of the energy analyser, the possible use of continuous ion sources (e.g. CI and fast atom bombardment (FAB)), and the possibility that MS-MS type information may be obtained by operating the electric sector and TOF analysers independently. A major disadvantage is the loss of sensitivity. This may well be overcome by use of an integrating transient recorder [29,30]. The ion optics of a TOFMS with multiple symmetry have been studied by Sakurai et al. [31]. Situations where the TOFMS satisfies the multiple focusing conditions (triple isochronous and triple space focusing) were investigated by introducing a symmetric nature into the arrangement of sector electric fields. Second-order characteristics were taken into consideration. High performance systems consisting of two or four electric fields were proposed. On the basis of this work, Sakurai et al. [32] subsequently published an account of a new TOFMS consisting of four 269’ electric sector fields in a clover-leaf formation. A flight path of 1.727m is achieved within a vacuum chamber 41 cm in diameter. TOF mass spectra of xenon, obtained using a 15ns (FWHM) pulsed electronic beam, indicated that a resolving power of 730 was obtained. The time spread of the mass peak due to transit through the analyser was estimated as 1 ns. This indicates that first-order isochronous focusing was achieved and that higher order aberrations were reasonably small. Two new approaches to the operation of a TOFMS have been suggested in a theoretical study by Reiss [33]. The first is a beam compression method which allows separation of one kind of particle (single m/z ratio) at a time, from a beam continuously emitted by an ion source. Owing to the continuous operation of the source a corresponding increase in signal-to-noise ratio is expected. The second method is based on the formation of a multiplex beam by modulating a continuously emitted ion beam. The use of a time array detector enables data to be collected. The algorithm of analysis is presented. This method should have an increased throughput advantage although an improvement in signal-to-noise ratio need not take place.

NEW TYPES OF TOFMS

Threshold photoelectron-photoion

coincidence TOFMS

(TPEPICO-MS)

The main restriction on TPEPICO-MS experiments has been the poor mass resolution (typically less than 30) of the TOFMS due to the low, i.e. 5-20Vcm-‘, extraction field applied to the ion source. A high field would reduce the energy resolution of threshold electrons. Nishimura et al. [34] have designed a new TOFMS for such work. This utilises a double-field ion source [5] and an ion reflector [35]. A resolution greater than 100 is achieved at low extraction field. TOFMS designed for exploration of the cometary dust composition

In 1986 the comet Halley entered the inner solar system. The European Space Agency’s GIOTTO mission spacecraft carried several instruments for analysing the composition of the dust forming the comet’s tail 1361. One of these instruments was a particle impact analyser (PIA) whose operation was based on the instantaneous ejection of atomic and molecular ions from a surface of a solid material during an impact of a fast dust particle. These ions can be detected and analysed by a specially designed TOFMS. The start signal for each mass spectrum can be achieved either by the light flash which accompanies the impact or by the charge pulse at the target. This signal initiates a (time)* function in order to achieve a linear mass scale. The flight path incorporates a reflector to enhance resolution. The mass range is l110 Da with a resolution of 150 at 110 Da. Up to 500 events per second can be processed although the transmission rate via telemetry is only 10 s-l. Therefore a means of selection of the spectra to be transmitted had to be designed into the control system. A study of the factors affecting the performance of a linear TOFMS modified for use with a surface ionisation source has been described by Chatfield and Ajami [37]. The long-term objective of these workers is the development of a technique to analyse the composition of particulate material for selected organic contaminants. Fourier transform TOFMS

Despite its early application in the field [38], the TOFMS has never really competed with the small quadrupole mass spectrometer as a sensitive low resolution detector for GC-MS instruments. The advantages of a low cost analyser and fast scan rate of the TOFMS fail to offset the reputation for low sensitivity. In an attempt to increase the sensitivity of the TOFMS for GCMS work, Knorr et al. [39] have recently reported some preliminary developments of a Fourier transform (FT) mode of operation. In theory the FT mode

8

of operation is capable of increasing the average ion current at the detector by a factor of 25 with no apparent loss in resolution. The FT operation cannot improve the fundamental resolution of the instrument. There are many analogies between FT TOF mass spectrometry and other FT techniques [40]. The normal modegate pulsing sequences are analogous to the entrance and exit slits of a dispersive spectrometer. Thus the trade-off between resolution and sensitivity intrinsic to all dispersive techniques may be anticipated. ET TOF mass spectrometry achieves the transform multiplex advantage through simultaneous detection of all ion flight times and the transform throughput advantage through essentially continuous source broadcasting. The operation of the technique was demonstrated by measuring the transformed electron impact (El) mass spectrum of toluene which was compared with that obtained in the normal scanning mode. A CVC Model 2001 was adapted for FT operation. Whilst the anticipated increase in sensitivity was not realised during FT operation, this was thought to be due to limitations in the equipment. The experiments did, however, indicate that the basic approach to FT TOF mass spectrometry is feasible and also the problems which need to be overcome to achieve success. MS-TOFMS

operation

The ability to identify relationships between parent and daughter ions which result from unimolecular decay or from collision-activated dissociation (CAD) or collision-induced dissociation (CID) is of increasing importance. Such information is obtained via the MS-MS technique, and the MS-MS spectra can be invaluable in mixture analysis and structure elucidation [41,42]. The majority of MS-MS work has been performed with instruments based on various combinations of magnetic (B) and electric (E) sector units and quadrupole (Q) filters. The sequential nature of the mass selection operations, however, allows examination of only one combination of parent ion and daughter ion masses at a time. Full MS-MS analysis is available only for samples that can be continuously introduced into the ion source over a relatively long time. The advantages of using time-resolved techniques to reduce the time required for MS-MS analysis have become apparent. Recently, two MS-MS instruments incorporating TOF have been reported. Stults et al. [43,44] have utilised ion beam pulsing and time-resolved (T) detection techniques in magnetic sector (B) mass spectrometry which they have termed time-resolved ion momentum spectrometry (TRIMS). This permits simultaneous momentum and velocity analysis of the ions providing energy-independent ion mass assignments. In principle, MS-MS spectra of all the ions in a mass spectrum should be acquired in a single sweep of the magnet, yielding significant improvement in scan rate or signal-to-noise ratio.

9

Colllslon Cell

Fig. 2. Schematic diagram of the ion momentum and time dispersion characteristics of a tandem magnetic/TOF (BT) instrument with post-sector beam deflection. (From ref. 46, Fig. 1.)

Details of the techniques developed for data acquisition and instrument control have been published [44]. Several accounts of work performed using this magnetic/TOFMS (BT) instrument have been published [45]. In the original instrument, ion source pulsing was used to facilitate TOF measurements. This resulted in severe limitations on the overall resolving power of the instrument. Subsequent development of a post-sector beam deflection technique has eliminated such limitations [46]. A schematic diagram of the experiment is shown in Fig. 2. With post-sector beam deflection, the ion source is operated under the conditions and configuration of a normal magnetic instrument. This gives the advantage of being able to operate at maximum magnetic field operation and sensitivity under normal focusing conditions. Thus the magnetic field and TOF features of TRIMS can be optimised independently. Whilst the BT technique is recognised as a valuable development in the MS-MS arena, there are some limitations to this configuration, e.g. the magnet must be scanned to obtain any MS-MS spectrum, and parent ion resolution is a function of kinetic energy release. This has led Glish and Goeringer [47] to develop the quadrupole/TOFMS (QT) shown in Fig. 3. The sample is introduced on a thermal desorption probe operated in the range 0-50V. Ions from the probe are focused by the source into the quadrupole which selects the required parent ions. The latter are focused by the einzel lens into a 10mm collision cell. Ions exiting the collision cell are

10

10 MM COLUSION CELL

TOF DEFLECTION ELECTRODE

Fig. 3. Schematic diagram of the tandem quadrupole/TOF (QT) instrument and acceleration scheme for positive ions. (From ref. 47, Fig. 1. Reprinted with permission of the authors and Anal. Chem. 0 1984, American Chemical Society.)

then accelerated by a four-element lens into a 50cm long drift region which is electrically floated at the accelerating potential of - 700 to - 1000 V for positive ions. A set of deflectors is mounted on the last accelerating lens to gate the ions into the drift tube for TOF analysis. The normal gate width used was 500ns. The multichannel analyser used to record the data is operated in a pulse-height analysis mode, so that the data are collected as flight time (mass) vs. abundance. In this configuration, only one ion per TOF pulse could be detected. This limitation was not serious since at the ion current levels used in this study, the probability of having more than one ion per pulse was extremely low. Conventional mass spectra were obtained by operating the quadrupole in the r.f. only mode, where it passes all the ions, and using the TOF analyser, or by scanning the quadrupole to select parent ions sequentially and gating the TOF analyser continually on to obtain daughter ion spectra. MS-MS studies of tetraethylammonium bromide indicate the applicability of the technique. In particular, the ability of QT to obtain MS-MS spectra sufficiently rapidly for the analysis of transient events was demonstrated. Possible improvements to the QT technique were also discussed. The relative merits of the QT and BT configurations are collated in Table 2. The major advantage of QT is the ability to acquire daughter ion spectra

11 TABLE 2 Comparison

Daughter ion spectra CAD energy Parent ion resolution

between QT and BT configurations

QT

BT

Not scanned

Magnet scanned

Low Quad

High Function of kinetic energy release during fragmentation

without scanning the quadrupole. Thus for analyses where only a few daughter ion spectra are desired the QT will have a time advantage over the BT. Also, parent ion resolution is determined solely by the quadrupole whereas for the BT configuration it is a function of the kinetic energy release. The other difference between the two approaches is that the QT operates at low CAD energies. This results in increased efficiency in fragmentation and collection. On the other hand, high energy CAD, accessible to BT, is much less sensitive to changes in collision gas pressure. The BT approach also has the larger mass range. The QT can operate as a normal MS-MS instrument; however, its fast operation does give it advantages over the scanning-type instruments for certain applications, for instance, pulsed ionisation techniques such as laser desorption. Another potentially valuable area could prove to be GC-MSMS. Owing to the scanning time necessary to obtain a complete MS-MS data matrix on conventional MS-MS instruments, the use of the latter for GC detection has been limited. Since the QT greatly reduces the time necessary to obtain a spectrum, GC-QT may become feasible in the manner originally postulated as a motivating factor for the development of the BT instrument [28,29,43]. A tandem TOF-TOF mass spectrometer has been constructed by Cooks and co-workers [48] for the study of polyatomic ion-surface collision phenomena. Ions are generated by EI and subjected to pulsed extraction, primary ion selection being by pulsed deflection after a 40 cm flight distance. Product ions are accelerated from the target surface and analysed by measurement of their total flight time. Daughter spectra of mass-selected parent ions compare well with surface-induced dissociation data from other instruments. A feature of this instrument is that the range of collision energies extends to above 300 eV. Internal energies as high as 30 eV can be deposited in selected ions by this procedure, so facilitating fragmentation and hence structural analysis.

12 SOLID-SURFACE

ANALYSIS

Plasma desorption mass spectrometry (PDMS)

A major stimulus for the resurgence of TOF mass spectrometry was the development of the 252Cf PDMS by Macfarlane [49,50]. The technique exploits high energy fission fragments from a 252Cfsource to bombard solid samples. Secondary ions ejected from the sample are analysed using TOF mass spectrometry which, because of its unlimited mass range, has enabled ions of mass in excess of 25000 to be detected. Fission fragments are also utilised to initiate the timing circuitry for each ionisation event, thus enabling very accurate (+ less than 1 ns) measurement of ion flight times. The initial interest in PDMS was, therefore, as an alternative technique to field desorption for the study of thermally labile molecules, particularly those of high mass, e.g. proteins. The technique has now been developed to the stage where it can be used for routine analysis of complex biomolecules not amenable to most of the standard mass spectrometric methods. Work has also been directed to the study of the decay of metastable ions within the instrument as well as to the investigation of the plasma desorption ionisation processes themselves. Future developments of the technique should be accelerated and enhanced by the appearance of the first commercial PDMS instrument [51]. Additional innovations include an instrument capable of analysing both liquid and thick solid samples [52] and a particle detector based on the detection of secondary electrons ejected by the impact of large organic ions [53]. The latter has been utilised to achieve postacceleration, yielding enhanced detection efficiency and identification of metastable decay channels. Geno and Macfarlane [54] have constructed a PDMS instrument utilising a particle guide consisting of two concentric cylinders that electrostatically capture ions in a spiral orbit as the ions traverse the flight path. The object of this work is to develop a TOF technique with low accelerating voltages (e.g. 5 V) where problems associated with beam divergence need to be minimised without perturbing the TOF mass spectrum. A number of excellent reviews concerning PDMS have appeared [55]. Macfarlane [50] has provided a clear account of the approach to the electronic measurement of the ion flight times and their conversion to mass numbers. The stability of the mass scale is indicated by the fact that it can be calibrated from the flight times determined for the ‘H+ and 23Na+ ions and is accurate to masses in excess of 20 000 Da. Sundqvist and Macfarlane [56] have reviewed the historical development of 252CfPDMS, its achievements and have also speculated as to its future. The limitations and possibilities of the PDMS technique have been discussed by Sundqvist et al. [57] whilst Cotter [58] has

13

tnm

13310

MASS

Fig. 4. The mass distribution of (a) tiger snake phospholipase homologue, (b) siamensis neurotoxin, and (c) bovine insulin. The solid envelopes have inverted relative widths at half-height (M/AM) as indicated. (From ref. 57, Fig. 3.)

provided a wide-ranging account of its application to the study of biomolecular species. One of the main criticisms that the PDMS technique has met is that of the low resolution of the TOF mass spectrometry technique used for mass analysis. Resolving powers (M/AM(FWHM)) of 500-2000 have been claimed for the instruments used to date. Since the flight times can be measured to less than 1 ns, the limitation on the resolution must be due to other instrumental factors, e.g. misalignment of the sample grids and detectors. Della-Negra and Le Beyec [59] have introduced a Mamyrin-type reflectron to reduce the effect of the initial velocity spread of the plasma-desorbed secondary ions. A resolution of around 2500 for organic molecules was reported. Although a 500-2000 mass resolution would appear to be woefully inadequate for the study of molecules of molecular weight up to and in excess of 20000 the situation is not as bad as it first appears [57]. For these large molecules, the relative abundance of the 13C,D, “N and I80 isotopes becomes appreciable. Consequently, a large molecular ion is associated with an isotopically averaged mass deduced from a distribution of masses [60]. As can be seen from Fig. 4, for the bovine insulin molecule (c), this is a skew distribution

14

whilst for the phospholipase A, molecule (a) it is almost symmetrical. If the envelope of the distributions is considered, then the inverted relative width, M/AM, is found to be in the 1000-2000 region. Therefore, if a distribution average is required to extract a molecular weight, the current typical TOFMS resolving powers may be adequate. In a mixture of large molecules differing in isotopically averaged mass by only a few units, the isotopic distribution mixing will cause serious problems even if high resolution were available. The limited resolution of the TOF mass spectrometry technique may, therefore, not be a serious disadvantage, particularly if PDMS is used in the role of molecule identifier. Such a role is significantly advanced when PDMS is used in conjunction with enzyme mapping techniques [61,62]. The reflex 252CfPDMS apparatus of Della-Negra and Le Beyec has been utilised to develop a coincidence method for studying metastable ion decompositions [63]. Two simultaneous TOF measurements, for ions and neutrals, are recorded event by event using the same start signals. Thus correlated reflex spectra can be obtained by setting electronic time “windows” on the neutral TOF spectra. These windows correspond to masses which have decayed in flight. The charged fragments are then selected by their coincidence in the correlated spectra; thus in-flight decomposition may be identified. The method has proved to be advantageous for the elucidation and/or confirmation of structures of organic compounds, e.g. adenosine and guanosine. Although in this case a 252Cf source was used, this reflectron coincidence technique is, in principle, applicable to any ionisation/desorption source. Metastable fragmentation of ions produced via 252CfPD has also been studied by Chait and Field [64]. The positive bovine insulin species investigated included the protonated molecular ion, a doubly charged quasimolecular ion, the dimer ion, chain fragment ions, and ions which result in the intense continuum which underlies the discrete peaks observed in the PD mass spectrum. A high proportion of the secondary ions entering the flight tube undergo unimolecular fragmentation. Investigation of the temporal distribution of the flight tube fragmentation indicates that these are fast processes. Thus most metastable decompositions occur early in the flight time. A novel utilisation of the PDMS technique is for the study of microscale chemical reactions in very thin (i.e. submonolayers to about 10’ molecular layers) films of non-volatile organic solids [65]. For this work, Chait and Field have used both a conventional 252CfPD mass spectrometer and a pulsed ion bombardment TOFMS. First, the film is analysed via PDMS which can be considered to be an essentially non-destructive technique. The film is then exposed to the reagent(s) of interest, e.g. light, 03, complex vapours, which cause(s) reaction to occur. Subsequently, the reacted film is re-analysed to provide information as to the nature and extent of the reaction. For example, photolysis using visible light of a rhodamine B film in a wet oxygen

1.5

atmosphere was found to produce three photo-de-ethylation products. Analysis of the films after different exposure times can be used to gain an indication of reaction rates. The advantages of the technique are high sensitivity and surface specificity. Low yields of non-volatile products can be reliably detected. PDMS is an attractive technique for surface and microscopic characterisation. TOF mass spectrometric analysis allows simultaneous multimass determination of atomic and molecular species [66]. An extension of the technique to facilitate depth profiling has been achieved by combining *%ZJfPDMS with kiloelectronvolt ion sputtering [67]. The PDMS depth profile of a 4660 A layer of aluminium on silicon was corroborated by X-ray photoelectron spectroscopy (XPS) depth profiling and indicated an information depth for PDMS of l-3 monolayers. In PDMS depth profiling, the analytical signal is generated independently of the sputtering step. Consequently, low sputtering rates can be employed to enhance depth resolution without loss of sensitivity. Given this independence between the generation of analytical info~ation and sputtering, the shallow escape depth, and the complete information range, PDMS combined with sputter depth profiling fills a unique niche in the array of surface analysis techniques. The feasibility of PDMS for spatially resolved chemical analysis has been demonstrated using a collimated 84MeV 84Kr7+ beam to induce deso~tion on a small sample area (about 11 pm in diameter); successive sample areas were probed by moving the sample in steps of a few micrometres [68]. An alternative approach to this microprobe mode is microscopy. The possibility of combining PDMS with transmission electron microscopy (TEM) has been discussed by Schweickert et al. 1661.The idea is that TEM is used to locate each primary bombarding ion by the track it leaves in the thin sample, e.g. mica, whilst the TOF system identifies the desorbed species. Whilst actual experimental studies were not reported, the approach is intended to obtain “chemical vision” on the submicrometre scale. One interesting phenomenon which has been observed in the Orsay PDMS laboratory is that of spontaneous desorption. In a PDMS instrument, with the “‘Cf source completely retracted, a potential of 5-12 kV was applied to a metal target on the surface of which were deposited organic or inorganic compounds. Under these conditions, it was found that negative ions and electrons were simultaneously emitted. After traversing the flight path, both particles were detected by the same microchannel plate assembly. A multistop time digitiser was used to determine the mass-dependent difference in flight times between the electrons and the ions [69]. The ion emission was found to be dependent on a high potential difference between the target and the ion accelerating grid placed a few millimetres away. It has been shown that the spontaneous desorption of ions from the target is in fact a secondary ion

16

emission phenomenon in which charged particles generated at the grid surface hit the target [70]. This type of TOFMS is the simplest and most inexpensive means yet devised for ionisation and molecular weight determination of thermally labile and non-volatile compounds, since virtually no ion source is needed for the generation of the incident particles. Laser microprobe

Clarke has provided a good general introductory review of the laser microprobe technique with an anticipation of future developments [71]. The LAMMA [72] and LIMA [73] instruments are now well established in the market place and many publications have appeared concerning their applications [74]. Potential distribution and ion trajectory calculations influencing the time development of ion motion in a laser microprobe have been performed by Vertes et al. [75]. Their results indicate that optimisation of geometric parameters should lead to more compact instrument design. Further calculations have been concerned with the non-linear optimisation of cylindrical electrostatic lenses [76]. Laser microprobe instruments provide rapid sensitive elemental, chemical and isotopic microanalysis with high spatial resolution. However, at best the technique can only be described as semiquantitative and attempts to improve on this have met with only limited success. A major barrier to quantitative analysis is that the established method of using a single laser for combined evaporation and ionisation is indiscriminate. Also, this approach results in less than 1% of the species desorbed being ionised. If the vast excess of desorbed neutrals could be ionised then the quantitative aspects of the technique would be greatly improved. Attempts are being made, therefore, to dissociate the desorption and ionisation steps by the introduction of a second ionisation laser [71,77,78]. One interesting modification of the LAMMA system is the addition of an optional straight flight tube and electron multiplier at the back of the ion reflector [79]. Any neutral species generated in the first flight tube pass through the reflector and are detected. Thus both ions and neutrals originating from metastable decay in the first flight tube may be simultaneously detected. An ambient pressure technique has been utilised to obtain mass spectra from pesticide residues on plant surfaces [80]. An automated LAMMA 1000 programmed to produce ion maps with high lateral resolution has been described by Wilk and Hercules [81]. Examples are given of the application of the technique to both organic and inorganic surface materials. The determination of the composition and energy distribution of laser desorbed products via a series of experiments is handicapped by the effect of pulse-to-pulse inhomogeneities in space and time of the laser beam and also

17

by unmonitored changes in the surface state produced by a series of pulses. Lazneva and Turiev [82] have attempted to circumvent these problems by developing a triple TOFMS for the simultaneous analysis of both positive and negative ions and also neutrals. This enables the required desorption information to be obtained for a single laser pulse desorption experiment. Applicability of the technique is indicated by a study of a Cd/O, surface. Although the laser microprobe had its original success mainly with inorganic problems, the laser desorption/ionisation technique has been applied to characterise thermally labile organic species [83]. Conzemius and co-workers have demonstrated the use of a subnanosecond pulsed laser to produce cationised parent ions as well as structurally significant fragment ions [84]. Details of their data systems, developed to measure low ion current pulses have been published [85]. The modification of a traditional Wiley-McLaren instrument for laser desorption experiments has been described by Cotter and co-workers [86]. Secondary ion mass spectrometry (SIMS)

SIMS is a successful analytical technique applied in surface, thin film and bulk analysis of solids. In more recent years it has been used to study biomolecules, polymers, pharmaceuticals and other involatile, thermally labile molecules. In this latter concern there are two main approaches: combination of a liquid matrix with a high resolution instrument (FAB) and combination of a solid matrix with a high transmission and low resolution instrument such as a quadrupole or TOFMS (static SIMS). FAB is characterised by high mass resolution and large sample consumption (greater than 10-‘Omol) whilst static SIMS requires extremely small samples (10-‘210-‘5mol) but has relatively low mass resolution. The advantages of the TOFMS over the more established quadrupole instrument in this application are the greater than lOO-fold improvement in sensitivity, no mass discrimination, simultaneous detection of the whole mass spectrum and the pulsed ion source. The pulsed ion source results in less damage being done to the bombarded surface and also a reduction or elimination of surface charge build-up which is a major problem in the case of non-conducting materials such as polymers. The application of TOF mass spectrometry to SIMS has been pioneered by Benninghoven at Munster and by Standing at Manitoba. The first SIMSTOFMS described by Benninghoven and co-workers [87] used a pulsed beam of argon ions to bombard a target with approximately 50 fmol mmm2 sample on the surface. The secondary ions are mass analysed via a TOFMS based on the original design of Poschenrieder [88,89]. This provides for second-order

18

energy focusing via an electrostatic sector field, together with two conventional linear flight tubes. The detector operates in the single-ion counting mode, giving increased dynamic range and very little mass discrimination at lower masses. The detectable mass range was increased by application to the conversion electrode of a post-acceleration voltage of up to 20 kV. Owing to the high sensitivity achieved, a mass spectrum can be obtained in approximately 10 s and involves the loss of only about 1% of the sample, typically around 1 fmol. The success of Benninghoven’s work has led to a commercial instrument of similar design [90]. This can also be used for laser microprobe work [91]. The Poschenrieder-type SIMS-TOF instrument had a typical mass resolution of 500 at a primary beam acceptance angle of 2”. As the resolution is limited by the second-order angular aberrations, an increase is only possible by reducing the acceptance angle. Eccles and Vickerman [92] used this approach to achieve a resolution of up to 2000 but with a considerable loss in sensitivity. In order to overcome this limitation in mass resolution due to second-order flight time aberrations of the Poschenrieder analyser, the Benninghoven group have developed a second version of the SIMS-TOF technique [93,94]. This is shown schematically in Fig. 5. Second-order energy focusing in the flight path of the secondary ions is achieved by a two-stage reflectron. A new electrodynamic mass separation and beam chopping technique based on a pulsed 90” deflection provides a primary ion beam of pulse width 1.5 ns. A mass resolution of 13 000 at 372 Da and a dynamic range of about six orders of magnitude have been obtained with this instrument. The instrument has been used to determine the chemical composition of the secondary ions directly from the spectra and to resolve peaks of oxygen- and non-oxygen-containing ions with AM = 0.030 at masses well above 200 Da. This information makes it possible to determine structures of ions originating from surfaces, which increases insight into secondary ion formation processes ]941. The original Chait and Standing SIMS-TOFMS [95] was described in our previous review [ 11.Recently, a 45” electrostatic mirror has been incorporated in front of the TOFMS detector [96,97]. The mirror deflects ions from a metastable decay through 90” to an ion detector whilst neutral species resulting from the same decay pass through the mirror to the normal TOFMS detector. Since the fragment ions have less momentum (same velocity, lower mass) than the parent ions, less time is required to deflect them in the mirror. Consequently they have a mass-dependent difference in arrival time at the ion detector. The parent ion is coincident with the detection of the neutral species which suffers no restraint on passage through the mirror (Fig. 6). The mirror and both detectors are mounted on a movable assembly to enable measurement of the ratio of the decay products to precursor ions at a number of drift distances and thus to determine the decay rate constant. Provided that a stable

19

(1) primary

ion Source

(2)

90’-defl*ctaf

(3)

bunching

(4)

maa8

reporation

(5)

targot

corouaJ

(6)

EiizeI-lena

(7)

two atop*

(6)

channelplate

(9)

photomultiplier

---- --

artem Jit

reflrctor and rcinl

-

Fig. 5. Schematic diagram of SIMS-TOF instrument utilising a two-stage reflectron. (From ref. 93, Fig. 1. Reprinted with permission of the authors.)

ion in the spectrum is used for normalisation, then the ratio measurement is independent of instrument transmission and detector efficiency, neither of which is likely to be identical for both parent and decay product. This group has also reported the development of a fast data system which enables complete spectra to be recorded at a total counting rate of 20 kHz. Such rapid scanning would be necessary if the SIMS-TOFMS were to be directly coupled to GC-MS or LC-MS. An off-line technique for collecting high performance

Fig. 6. Effect of the electrostatic mirror on the TOF spectrum of the tripeptide glycylglycylphenylalanine molecular ions. The upper spectrum is the normal spectrum with the mirror off. When voltage is applied to the mirror, the neutral daughters travel to the 0’ detector (bottom spectrum) while the charged daughters and undissociated parent ions appear in the spectrum from the 90’ detector. (From ref. 97, Fig. 3.)

liquid chromatography (HPLC) effluents for subsequent SIMS-TOFMS analysis has also been developed [98]. A detailed description of the design and development of a new improved instrument has recently been published [97]. A single-stage ion mirror results in a resolving power of 10000 whilst the signal-to-background ratio is also much improved. Two detectors are used: one situated behind the mirror is used to detect neutral species from unimolecular decay in the first linear flight path; the second detector measures the parent and daughter ions in the reflected spectrum. This system enables correlations between the charged and neutral daughters to be determined, thus yielding information about the structure of the parent ion. This coincidence approach is similar to that of Della-Negra and Le Beyec [63] discussed under PDMS. Cotter and co-workers [99,100] have reported the development of a liquid secondary ion-TOFMS instrument configured by utilising a Kratos Minibeam I ion gun as a source of the primary ion beam and a CVC Model 2000 TOFMS for secondary ion analysis. The field-free ion source is well adapted to the use of a liquid matrix, and the electron grid pulse from the TOFMS electronics is used to control the pulsing of the ion beam. Time delays between ion formation and extraction from the source region improve focus-

21

ing and allow observation of fragments from metastable decompositions. The molecular yield is found to be dependent on sample concentration, instantaneous primary ion current and the primary pulse repetition rate [loll. The dependence on primary pulse repetition rate indicated that a finite recovery time is required to repair radiation damage from high flux particle beams. For many biochemical analyses for which better than unit mass resolution is not required, the technique may offer a low cost alternative to conventional liquid SIMS for molecular weight and sequence ion determination. A continuous flow probe interface to enable the technique to be coupled to an HPLC apparatus has been evaluated [102]. The SIMS technique has considerable dynamic range and in favourable cases can detect certain elements to a few parts per billion. Sensitivities below this level are seldom achieved. This detection limit arises from a number of sources. Firstly, as was also indicated in the case of the laser microprobe, only. about 1% of the sputtered material is ionised; the remainder, being neutral, is not detected in the mass spectrometer. Secondly, the magnetic and quadrupole instruments used in a typical SIMS apparatus have ion transmissions in the range 10-‘-10-4. The final problem is that secondary ion formation is strongly influenced by electronic effects arising in the sample matrix, making it extremely difficult to quantify most measurements. Obviously, a method of analysing efficiently the abundant neutrals ejected from an ionbombarded surface would be of major benefit to the SIMS technique and there is considerable interest in developing such an approach. Kimock et al. have coupled multiphoton resonance ionisation (MPRI) to energetic ion bombardment to yield a highly efficient and selective tool for solids analysis [103]. There are a number of major advantages to this approach. All elements except helium and neon can be detected. Every neutral species passing through the laser beam can be ionised; thus MPRI samples the majority of the ejected particles. Examination of the neutrals should greatly reduce matrix effects and thus improve prospects for quantitative analysis. MPRI can be made selective to a single element by choice of excitation wavelength. This would, for example, easily distinguish between P+ and SiH+, a problem which has beset the SIMS analysis of phosphorus-doped polysilicon semiconductors [ 1041.This selectivity obviates the requirement for a high resolution mass spectrometer. Thus the TOFMS with its ability to analyse all ions simultaneously has a considerable advantage for the analysis of the MPRI species. Although this technique has high promise there are a number of experimental factors which may ultimately limit the sensitivity. Kato et al. [105] have calculated the mass resolution for a TOFMS with a two-stage reflectron, applied to secondary neutral mass spectrometry. Instrumental parameters are optimised for energy and space focusing, correcting the flight time difference due to the energy width AE of the sputtered

22

particles and the spatial width As of an ionising laser beam. The effect of As can be compensated by applying an accelerating field to the ionising region, and the maximum resolution becomes about 1000 for AE = 10 eV and A.r = l.Omm. Utilisation of non-resonant multiphoton ionisation of secondary neutrals prior to TOF analysis has been proposed by Bentz and Honig [ 1061in a design for an organic SIMS instrument with a separate triple-stage quadrupole. The latter will provide structural information up to its mass limit of around 1000 whilst the TOF system will enable access to be made to a much higher mass range. The TOF-SIMS instruments discussed to date are somewhat complex. However, many basic aspects of secondary-ion production can be explored using a simple linear configuration. Since this can be very compact the approach is ideally suited for yield measurements at different angles of primary ion impact. Szmczak and Wittmaack [ 1071have described the design and performance of a TOFMS with variable tilt angle for sputtering studies in the nuclear stopping regime. An extension to the SIMS approach to provide a general method of surface analysis by laser ionisation (SALI) has been described by Becker and Gillen [108,109]. A small fraction of a monolayer is desorbed or sputtered from a surface by a probe beam (Ar+ ions, laser, electrons). The more abundant desorbed neutral fraction is ionised above the surface by non-resonant multiphoton ionisation and the photoions are then analysed by a reflecting TOFMS with mass resolution of about 1000. A major advantage which SAL1 provides is that desorption and ionisation are separated and can thus be independently controlled. Matrix effects are greatly reduced so that the relative variation of the neutral yields as a function of local chemical composition is very small, in contrast with the yields of secondary ions. Photoionisation via the high power untuned UV laser often results in saturation of the non-linear multiphoton ionisation process for all species present in the laser local volume. This is important for correct determination of stoichiometries. The choice of non-resonant multiphoton ionisation and the factors limiting absolute quantitation of the method have been discussed [l lo]. Application to a wide range of surface and material analyses has shown the versatility of the technique which can operate to the 1 ppm level without extensive signal averaging and rare gas ion beam currents of about 1 PA. Extension of the technique to depth profiling has been reported [l 111. Use of a more powerful primary desorption beam, 1 A cm-* in a 0.2pm diameter beam, will allow development of the technique to obtain submicron lateral resolution and also its use for three-dimensional imaging [l 111. Molecule microscopy depends on the spatial variations of emissions of neutral molecules from a sample and produces images that reveal variations

23

Fig. 7. Sketch indicating mode of operation of proposed electron stimulated desorption scanning molecule microscope. (From ref. 112.)

in the number of molecules present on or emerging through the surface of the sample. These molecules can have been part of the sample or applied as a surface stain. King [112] has described the design of a proposed electron stimulated desorption scanning molecule microscope (ESDSMM) which offers the possibility for nanometre resolution across the surface. A schematic representation of the proposed technique is presented in Fig. 7. The sample, located on a substrate 10 nm thick, would be subjected to 1 ps desorbing electron pulses. The desorbed neutral species will impinge and stick on the cold (77 K) tungsten tip of radius 1 pm. This tip would oscillate from a position of 1 pm above the chosen location on the sample at the time of the

24

electron pulse to a position above the entrance to a TOFMS between such pulses. Above the TOFMS a 1 ns 900V pulse causes field ionisation of the desorbed species. The resultant ions then enter the TOFMS for analysis. Provided the ambient pressure in the system is maintained below lop9 Pa, less than one molecule will strike the tip per desorption/detection cycle. The advantage of the TOFMS for this application is that the complete mass range is monitored for each cycle and it exhibits high efficiency for single-ion transmission and detection. Atom probe Atom probe techniques [113] are based on field evaporating atoms from a specimen surface using high voltage pulses of short duration and a fast rise time. These pulses also trigger a timing chain which is interfaced to a detector and other discriminating electronics. In the original design [114] a specimen goniometer was used to bring a feature of interest under a probe hole cut in a channel plate intensifier system. High voltages were then applied, the probe hole becoming an aperture into the drift tube of a TOFMS for analysis of the ejected ions. Considerable interest is being shown in the development of the TOF mass spectrometry aspects of this technique. Tsong [115-l 181 has presented methods for achieving a better accuracy in the ion energy analysis and time resolution in ion reaction time measurement. Sakurai et al. [I 191 have described a high performance, focusing-type, TOF atom probe with a channeltron as a signal detector. A TOF atom probe designed to examine radiation defects in metals has been reported by Bobkov et al. [120]. Reinmuller [121] has described an automatic control and timing unit for TOFMS used in an atom probe or field desorption system. An extension of the conventional atom probe is the use of a short-duration laser pulse and a high electric field to generate ions at a solid interface. This pulsed-laser atom probe can be used to probe the composition and structure of a sample with true atomic, spatial and depth resolution. Independent adjustment of the electric field can be utilised to study surface chemical reaction. For a detailed account of the many advantages and applications of this technique, the reader is referred to a review by Kellog [122]. PULSED

LASER IONISATION

Lasers have found increasing applications in mass spectrometry for the desorption of non-volatile thermally labile molecules and ions, for gas phase photoionisation of neutrals and for the photo-fragmentation of neutrals or ions. TOF mass spectrometry with its high transmission and ability to provide

25

a complete mass spectrum for analysis per ionisation event has been extensively used in this area. Laser desorption mass spectrometry for non-volatile organic molecules has been reviewed by Cotter and Tabet [83] and more recently by Hillenkamp [ 1231. Development of the technique requires a much greater understanding of the various desorption and ionisation processes occurring than is currently available; considerable effort to this end can be anticipated. Hillenkamp’s research group has modified a LAMMA instrument to enable laser desorption of neutrals to be followed by photoionisation with a second laser [124]. The experiment allows a detailed study of the parameters affecting both processes to be undertaken. Among the conclusions drawn are that the two-laser system, in which the two processes can be independently controlled, shows increased sensitivity and reduced fragmentation. The objective of such studies is the development of a model for the desorption/ionisation process. One such model has been proposed by Zare and Lavine [125] in terms of bond-selective processes in rapid desorption. A criterion for the required laser power is derived. Elemental and isotopic analysis

Resonance ionisation (RI) mass spectrometry is a technique currently being developed for elemental and isotopic analysis of solid samples [78]. As observed by Gross [ 1261the isotopic shifts in the atomic spectra are less than the Doppler line width for atomic numbers between 10 and 200 and at a pressure around 1 Torr. Under these circumstances, RI will ionise all the isotopes simultaneously. Provided the process is saturated for all the isotopes of the element under investigation, the ionised atoms will be an accurate isotopic representation of the sample. An RI-TOFMS system for elemental and isotopic analysis was first proposed by Beekman et al. in 1980 [127]. The applicability of the method was demonstrated by the detection of chromium in stainless steel [128]. A tunable dye laser was used to select the Cr atoms present and a mass spectrum of the four stable isotopes of chromium produced. Potential applications of the technique are discussed. Peuser et al. [129] have used laser photoionisation to detect trace amounts of plutonium. High sensitivity and selectivity was achieved via three-step excitation and ionisation of Pu atoms, at high pulse-repetition rates, followed by TOF mass spectrometry analysis. The laser system consisted of three tunable dye lasers simultaneously pumped by a copper vapour laser. Samples containing between 10” and 10” atoms of 239Pu on rhenium filaments were measured yielding strong resonance signals with maximum count rates of several hertz at a vanishingly low background. A detection efficiency of lo-’

26

was determined allowing detection of approximately lO*Pu atoms in a sample. RIMS techniques have also been applied to the isotopic analysis of neodymium [130], tantalum [131,132] and technetium [133]. Miller [134] has reported the ultrasensitive, isotopically selective analysis of nitric oxide in a supersonic beam. The extension of this work to “single-molecule” detection is discussed. A major difficulty in isotope ratio measurements by TOF mass spectrometry is precise quantification of the fast signal pulses. A fast-pulse detection system designed to mitigate this problem has been described by Green et al. [135]. Fassett et al. [136] have used time-resolved magnetic dispersion for large isotope ratio measurements in RIMS. Multiphoton ionisation (MPI) The potential of the MPI-TOF mass spectrometry technique, both for analysis and the investigation of RI spectra and ion decomposition processes, was discussed in a previous review [ 11.MPI mass spectrometry reveals characteristic features which are different from those typical for most conventional ionisation methods. Species selectivity, isomer specificity, soft ionisation and hard fragmentation are now well understood on the basis of the unique excitation mechanism in MPI mass spectrometry. Neusser [ 1371has reviewed the existing models for the excitation mechanism and discussed the conclusions to be drawn with respect to structural analysis and for the investigation of elementary reaction steps in ion kinetics. For MPI experiments conducted in the conventional Wiley-McLaren source instruments, Martin and O’Malley [138] have shown that the time-lag delay between ionisation and ion withdrawal into the flight tube is an important experimental parameter in the measurement of the fragmentation patterns obtained via TOF mass spectrometry. In order to be able to measure ionisation efficiencies over a particular class of compounds, Opsal and Reilly [139] utilised a GC-MPI-TOF mass spectrometry technique. A known mixture was injected into the gas chromatograph so that known quantities of the individual compounds were eluted into the ion source. Thus the mass spectra of the individual species could be obtained under identical conditions. A study of alkylbenzenes is reported. A TOFMS, which provides access to an entire photoionisation mass spectrum for each laser pulse, is central to a laser-based multidimensional analytical instrument devised for selective and sensitive on-line detection of polycyclic aromatic compounds [140]. The effluent of a capillary GC is interrogated by a tunable UV laser beam at collision-free pressures. Selective excitation/ionisation occurs, based on the spectroscopic absorption characteristics of the analyte molecules. The laser-analytic interaction products (cation, electrons and photons) are simultaneously

27

monitored, permitting all the analytically useful data to be extracted “on the fly”. Absolute detection limits in the low picogram range and a linear dynamic range of four decades are reported. Marijnissen et al. [141] have described a proposed multiple laser-TOFMS system for the analysis of size and chemical composition of individual aerosol particles “in situ” on a continuous and real time basis. Kuhlewind et al. [142] have reported on the MPI mass spectra produced from a homologous series of alkyl cations. The objective was to investigate the fragmentation pattern transferability from one ion, C,H&+, , to the next higher one. In electron impact ionisation all ions in the same mass range, even when produced from different parent species, produce similar fragmentation. In direct contrast, MPI mass spectra of homologous compounds display strong differences. The wavelength of the exciting light influences the fragmentation pattern in a direct way, thus enabling two-dimensional control of the fragmentation pattern. This can be exploited for mixture analysis. Welge and co-workers [143,144] have utilised a TOFMS to investigate two different types of photochemical process induced by laser radiation: (i) IR multiphoton excitation and dissociation of gas phase molecules and ions; (ii) UV-laser photoablation of polymer surfaces. The addition of a supersonic molecular beam sample cooling technique further facilitates selective ionisation by transforming the UV absorption spectrum of the sample, broad at room temperature, to one which exhibits sharp vibronic features under supersonic beam conditions. Trembreull and Lubman [145] have used this technique to investigate resonant-two-photon ionisation (R2PI) spectra of ortho, meta and para isomers of cresol to evaluate the method as a sensitive means of detecting and discriminating these compounds by monitoring the parent ions in a TOFMS. The laser powers used were chosen to avoid fragmentation. The possibility of direct atmospheric air sampling was investigated and an estimate of the real discrimination (better than 1:300-500) and sensitivity limits (about 20 ppb) are discussed. A three-photon ionisation technique has been utilised for the detection of azabenzenes whose ionisation potentials are too high for R2PI [ 1461.Syage et al. [147] have investigated the sensitivity and selectivity of a supersonic beamMPI-TOFMS for atmospheric monitoring. Lubman and Jordan [I481 describe an improved design of a TOFMS constructed for optimum use with resonance-enhanced MPI (REMPI) in supersonic beams. The design utilises fast-pulsed molecular beam techniques to allow sample reservoir pressures above atmospheric and a large sample exit orifice (0.5 mm) in order to provide high on-axis intensity for maximisation of the photoionisation with the use of only modest pumping capacity. The supersonic beam is sampled by a skimmer before entry into the laser ionisation region. Differential pumping and a liquid-nitrogen-cooled baffle maintain the ionisation/acceleration region and flight path below 5 x lo-‘Torr.

28

An unusual feature is that the TOFMS is located vertically, saving laboratory floor space and facilitating construction of the differential pumping and cooling of the ionisation region. Voorhees and Seliskar [149] describe a versatile trigger and time-delay generator for this type of instrument. In general, the seeded supersonic molecular beam is orthogonal to the flight path of the ions. Thus the ions will have an initial velocity component perpendicular to the flight path. This will cause the ions to deviate from a straight line path to the detector and hence a major percentage of ions miss the detector unless some means of compensation is introduced. The simplest method is the use of deflection plates immediately after the ion source [148]. Conover et al. [150] have recently shown that by suitably tilting the acceleration grids in the ion source, the initial velocities of molecular cluster ions transverse to the flight path can be selectively reduced to zero, allowing for a less-off-axis flight path and permitting detection of higher mass clusters than is possible with conventional post-acceleration deflection schemes. The alternative approach, which avoids this particular problem, is to direct the supersonic beam along the first flight path of a V-type reflectron. This has been successfully used in the LAMMA and LIMA instruments and also by the Grotemeyer group. The REMPI-TOF mass spectrometry technique has been shown to be applicable to species which are volatile below 150°C. However, the future may well show that its greatest utility is for the analysis of non-volatile and/or thermally labile biological and pharmaceutical molecules. It is, therefore, important to develop sampling methods which inject such molecules into the supersonic jet system. Lubman and co-workers have investigated the applicability of supercritical fluid injection [ 1511. Supercritical fluids, which have properties between those of an ultradense gas and a liquid, have been shown to act as solvents for polynuclear aromatic hydrocarbons and small thermally labile biological molecules. Supercritical fluid injection has been interfaced with mass spectrometry for the detection of species such as T-2 toxin and diacetoxyscirphenol [ 1521. Lubman and co-workers have developed a supercritical fluid injection system coupled to the laser REMPI-TOFMS system described earlier. Both carbon dioxide and nitrous oxide have been used as fluids up to at least 380 atm and 100°C. The high pressure expansion is accomplished with modest pumping requirements owing to the use of a specially designed pulsed, high pressure valve [153]. Applicability of the technique is illustrated by mass spectra obtained from carbazole, tyramine (m.p. 163°C) and tryptophan (m.p. 19OOC).Thermal decomposition of the last two compounds, which was a feature of the use of a heated inlet for admission into the original REMPI-TOFMS, was absent when supercritical fluid injection was used. The solute does not necessarily have to be dissolved in a supercritical fluid. In a subsequent article, Lubman and co-workers [154]

29

described the application of similar technology for the high pressure injection of solutions of thermally labile biological molecules, e.g. catecholamines and indoleamines, in methanol. This approach had the advantage that the excessive clustering that normally occurs in continuous jet expansion was not observed. Supercritical fluid injection, and the possible alternative thermospray technique [155] both have the major disadvantage that large amounts of solvent are injected into the mass spectrometer. A better method of introducing low volatility, thermally labile molecules into the REMPI region may well be pulsed laser desorption, followed by cooling in a supersonic expansion jet of inert gas. Desorption is normally from a solid sample although a potentially useful alternative would be the FAB technique of dissolving the sample in glycerol so that the sample remains homogeneous because of the fluidity of the matrix. Such an approach has been described by Li and Lubman [I 561 utilising a LD-MPI-TOFMS system described previously [ 1481571. Grotemeyer et al. [158,159] have shown that “double-barrelled” laser desorption-REMPI mass spectrometry has many advantages over the singlestep desorption/ionisation technique, particularly if combined with a reflectron TOFMS (resolving power in excess of 10 000 is possible [160]) for analysis. Large biomolecules can be vaporised intact. The extent of hard or soft ionisation, and hence the degree of ion fragmentation can be controlled via the ionisation laser intensity. This tunable degree of fragmentation provides a valuable tool for structural analysis of these biomolecules. Confusing ion formation, from alkali metal addition or proton addition or due to matrix effects, is minimal. The effectiveness of the technique for the study of large labile biomolecules is illustrated with reference to studies of chlorophylls, porphyrins and a peptide. Grotemeyer and co-workers have had considerable success with this technique and numerous papers concerning its application have been published, see for example, refs. 161 and 162. A commercial instrument based on this work is now available [163]. One difficulty with resonant laser ionisation is that the REMPI absorption spectra of many compounds are not widely known and so it is not known in advance if there is an appropriate wavelength for resonance ionisation. This is particularly a handicap in attempts to use the REMPI-TOFMS technique as an analyser of unknowns, e.g. as a detector for the new generation of high resolution chromatographic techniques. In this respect it would be highly advantageous to couple the MPI source with the general EI technique. EI would complement MPI by providing identi~cations of unknowns via the large library of EI spectra available or by interpretation of unknown EI spectra by well-established rules. Introduction of a conventional heated filament as a source of electrons requires pulsed voltages on the electron and ion accelerating grids. While this can be done successfully [164], it does remove

30

one of the advantages of the pulsed laser TOFMS, namely the use of static as opposed to pulsed voltages. A novel and simple way of avoiding this dilemma has been devised by Grotemeyer and co-workers [ 1651.They have shown that it is possible to generate very short, high intensity electron pulses via pulsed laser interaction with tantalum. In their system the 0.25 mm tantalum wire is located between the repeller and first ion extraction grids of the ion source and held at a potential intermediate between those on these two grids. The pulsed laser normally used for MPI is also used to produce the electron pulse via interaction with this tantalum wire. This system allows medium-resolution mass spectra to be recorded without pulsing the ion accelerating voltages in the TOFMS. The EI spectra reported show good correlation with those in the literature. An alternative approach to the identification of the unknowns would be to use nonresonant laser ionisation in the vacuum UV. Ionisation would be essentially single photon to produce mainly easily identifiable molecular ions [ 1661. A tandem TOFMS has been developed by Johnson et al. [167] to study the photodestruction cross-section for mass-selected (CO,): clusters. To avoid the complication involving internal excitation of ion clusters encountered when ionising neutral clusters, a method has been developed for synthesising the ion clusters by nucleating them onto the monomer ion via clustering reactions occurring during free-jet expansion. The ion clusters thus formed are allowed to drift some 1Ocm in a field-free region before being pulse ejected into the first flight tube for mass selection of the parent clusters. The TOF spectra thus have as a time origin the rapid switching of the draw-out voltage, not the usual laser-pulse-initiated TOF spectra where the ions are created in a constant field region. The clusters are photodissociated during the free flight by passing a pulsed laser beam down the axis of the flight tube. Parent ions are separated from daughters by a reflecting electric field and are detected at the end of a short free-flight region-the second TOF. Smalley and co-workers [168,169] have constructed an elegant tandem TOFMS to monitor the dissociation caused by pulsed-laser photolysis of initially mass-selected cluster ions. This spectrometer is optimised to allow for maximum overlap of the ions and dissociation laser beams as well as equal detection sensitivity for parent and daughter particles. The apparatus is specifically designed for the study of the photofragmentation of isolated, internally cold, chemically bonded cluster ions. The spectral utility of the technique is illustrated in the cases of Ni: and Nbl for which well-resolved vibronic bands are reported. For larger more complicated metal clusters the absorption spectrum is far too congested and perturbed for analysis, but photodissociation can reveal the thresholds and time scales of various fragmentation pathways. TOF mass spectrometry has been used to show that stable clusters consisting of 60 carbon atoms can be produced by laser

31

irradiation of graphite [ 1701. Such end-to-end tandem TOF experiments are difficult in that the fast ion beam must be overlapped in time and space with a nanosecond pulsed laser. Deceleration and re-acceleration of the ion beam can provide a partial solution to the problem but the timing is still critical. Duncan and co-workers [171] have described a novel reflectron tandem instrument which provides ion-photodissociation laser interaction to occur at the turnaround point in the reflectron, i.e. when the ions are essentially stationary. This provides optimum overlap of the laser and ion beams whilst minimising the uncertainties in the photodissociation laser firing time. Potential developments of this approach are discussed. A novel method for analysing delayed dissociation of large clusters of ions or heavy ions has been devised by Leisner et al. [ 1721who have combined a TOFMS with differential energy analysis. The main interest in LD-MPI-TOF mass spectrometry has been in the identification of complex molecules. As yet little attention has been paid to the quantitative aspects. With this in mind, Zare and co-workers have applied the technique to quantify surface adsorbed molecules [173]. The dependence of ion signal on sample concentration in the fresh area (5 x 10-4cm~2) irradiated by each desorption laser pulse was found to be linear from nanomoles to subfemtomoles, corresponding to a surface coverage of 102-lo3 monolayers. A detection limit of 4 x IO-“moles was obtained for protoporphyrin 1X dimethyl ester [174]. A significant problem associated with MPI is that the unusually high selectivity in ionisation probability for organic compounds leads to very non-uniform detection sensitivities; this places severe limits on the ability to quantify relative abundances in mixtures, particularly those containing unknowns. This selectivity tends to diminish as the wavelength of the light used for MPI moves into the UV region. Becker and co-workers [ 1661 have suggested single-photon ionisation using coherent vacuum UV light as a “universal” method of overcoming this problem. They illustrate the approach by the use of a laser pulse at 118 mm (10.5eV) in conjunction with their SAL1 technique discussed earlier. A similar approach has been taken by Huth and Denton who have used a UV hydrogen laser as a selective ion source for molecules which ionise below 7.8 eV [175]. Such compounds include many pharmaceuticals and drugs of abuse as cocaine, heroin, phenylcyclidine, methamphetamine and LSD. Only parent ions have been observed for the compounds studied. The selectivity of the threshold ionisation process is very high; compounds with ionisation potentials above 8.0eV are completely rejected. Photoactive species can be injected as solutions in organic solvents without mass spectral interference from the solvent.

32 HIGH MASS STUDIES

Determination of molecular weight is a fundamental measurement required for the elucidation of structures of biomolecules. It is only in the last ten years or so that ionisation techniques, e.g. field and plasma desorption, FAB, capable of yielding ions above 2 kDa have been developed, thus providing the possibility that mass spectrometry can be applied to large biomolecules. This development requires extension in the mass range of instruments and improvements in high mass ion detection. Because of this, the attribute of the TOFMS which is currently attracting the most attention is its unlimited mass range which gives it an inherent advantage in the drive to extend the accessible mass range. Major limitations on the mass range are the efficiency of ion production and detection. The high transmission capability of the TOFMS facilitates solution of the latter problem. An overview of high mass analysis has been provided by Gaskell [176], whilst Morris [177] has discussed strategies developed for mass spectrometric analysis of high mass biopolymers. Until recently, PDMS (the natural TOF mass spectrometry technique) has been most successful in studying large intact biomolecules such as those from ovalbumin (molecular weight 45 kDa) [178]. FAB and SIMS have also been successfully applied to the analysis of proteins and polymers [179,180]. The electrospray technique [181] is currently attracting attention as a means of generating highly charged, high molecular weight, protein ions. Because of the highly charged nature of these ions, their m/z values fall within the range of available multisector instruments capable of precise m/z measurements, with obvious advantages for empirical formula determination. In the last 2 years, LD-TOF mass spectrometry has raised the upper mass limit accessible to mass spectrometry to 200 kDa with the hint that even lo6 Da ions are obtainable. Tanaka and co-workers, at the second Japan-China Joint Symposium on Mass Spectrometry in 1987, reported the nitrogen-laser desorption of protein molecular ions up to 34 kDa. Their technique utilises the previously mentioned “gradient electric field ion reflector” of Yoshida [19] and has since provided spectra above 100 kDa [20]. Hillenkamp and coworkers [123,182,183] report the detection of ions at 200 kDa and above produced via UV laser desorption in a LAMMA 1000 instrument using post-acceleration to enhance detection of the slow moving heavy ions [184]. Both these groups use a matrix to absorb the UV light and to convert electronic excitation into vibrational or translational energy, expelling the ionised sample species into the gas phase. The essential difference between the two is the type of matrix. Tanaka et al. [20] use a liquid slurry of analyte and finely divided metal powder in glycerol whereas the Hillenkamp group [123,182,183] use a dried mixture of the sample dispersed in nicotinic acid.

33 TABLE 3 Commercially available TOFMS instruments Manufacturer

Type

Reference

cvc

Wiley-McLaren LAMMA LIMA

187 72 73

252CfPDMS TOFMS-SIMS LD-MPI-TOFMS LD-MPI-TOFMS Velocity compaction “LE-PD-ion beam” TOFMS

51 91, 92, 188 163 189 26 190

Leybold Cambridge Mass Spectrometry (Kratos) Bio-Ion VG Ionex Bruker Jordan Toftec Nermag

Beavis and Chait [185], using a simple linear TOFMS, have confirmed the production of high mass ions via laser desorption from a dried matrix. Four new matrix materials with properties as good as or better than nicotinic acid are reported. Sundqvist and co-workers, using a BIO-ION PDMS instrument adapted to facilitate laser desorption, have also been able to reproduce Hillenkamp’s observations [ 1861. Laser desorption has undoubtedly arrived as a technique for producing high mass ions. Attention is now being focused on identifying the factors which significantly influence the process. Compared with PDMS and FAB, it should be much easier to control experimental parameters, e.g. laser wavelength, intensity and pulse length, matrix, etc. in order to gain insight into and suggest models for the phenomena involved. The effect of instrumental parameters, particularly on the detected intensities of high mass ions, is also important [185,186]. CONCLUSIONS

The resurgence of interest in TOF mass spectrometry has been due to the advent of applications such as laser and plasma desorption, laser ionisation and surface analysis, which require its ability to provide a complete spectrum per ionisation event and also unlimited mass range. In order for this renaissance to continue and expand, the technique needs a wide base of commercially available instruments. It is heartening that, as can be seen from Table 3, such a base now exists. The companies concerned are to be congratulated on their efforts and the TOF mass spectrometry community wishes them a prosperous future.

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