Restricted Energy Transfer in Laser Desorption of High Molecular Weight Biomolecules

Scanning Microscopy Volume 5 Number 2 Article 3 4-20-1991 Restricted Energy Transfer in Laser Desorption of High Molecular Weight Biomolecules Akos Vertes University of Antwerp, Belgium, [email protected] Renaat Gijbels University of Antwerp, Belgium Follow this and additional works at: https://digitalcommons.usu.edu/microscopy Part of the Biology Commons Recommended Citation Vertes, Akos and Gijbels, Renaat (1991) "Restricted Energy Transfer in Laser Desorption of High Molecular Weight Biomolecules," Scanning Microscopy: Vol. 5 : No. 2 , Article 3. Available at: https://digitalcommons.usu.edu/microscopy/vol5/iss2/3 This Article is brought to you for free and open access by the Western Dairy Center at DigitalCommons@USU. It has been accepted for inclusion in Scanning Microscopy by an authorized administrator of DigitalCommons@USU. For more information, please contact [email protected]. Scanning Microscopy, Vol. 5, No. 2, 1991 (Pages 317-328) 0891-7035/91$3.00+ .00 Scanning Microscopy International, Chicago (AMF O'Hare), IL 60666 USA RESTRICTED ENERGY TRANSFER IN LASER DESORPTION OF HIGH MOLECULAR WEIGHT BIOMOLECULES Akos Vertes* and Renaat Gijb els Departm ent of Chemistry, University of Antwerp (U.I.A.), Universiteitsplein 1, B-2610 Wilrijk (Belgium) (Received for publication November 15, 1990, and in revised form April 20, 1991) Abstract Introdu ction Producing ions from large molecules is of distin With the growing importan ce of biomedical investi guished importance in mass spectrometry. In our present gations in organic analysis the emphasis has been shift study we survey different laser desorption methods in ing to the detection and structure determination of ever view of their virtues and drawbacks in volatilization and larg er and more complex molecules. Gas phase analyt ion generation. Laser induced thermal desorption and ical methods in general - and mass spectrometry was matrix assisted laser desorption are assessed with spe not an exception - exhibited increasing handicap with cial emphasis to the recent breakthrough in the field the growing molecular masses, because of the involatility (m/z > 100,000 ions produced by matrix assisted laser and instability of these materials. desorption). Efforts to understand and describe laser de The early answer to this challenge came in the sev sorption and ionization are also reported. We emphasize ent ies, in the form of the so called "soft" ionization tech the role of restricted energy transfer pathways as a pos niques. Field desorption, chemical ionization, plasma de sible explanation to the volatilization of non-degraded sorption, secondary ion, electrohydrodynamic and laser large molecules. desorption ion sources were developed to cope with non volatile compounds. A thorough overview of these meth ods with respect to their high mass capabilities was pub lished by Daves in 1979. Th ere seemed to be two dis tinct strategies to follow: (a) optimization of volatiliza tion conditions in terms of sample dispersion and heating rate and (b) direct ionization of molecules from a surface by exte rnal electrostatic field, particle bombardment or laser radiation . At this stage of development, however, analytical possibilities were limited to the mass range m/z < 10,000. During the eighties four techniques were able to overcome this limit. 252 Cf plasma desorption mass spectrometry (Sundqvist and Macfarlane, 1985) became available in the m/z < 45,000 mass range. Fast atom bombardment - an offspring of secondary ionization with spec ial sample preparation - reached the m/z ~ 24,000 region (Barber and Green, 1987). The two fore runners, however, became the electrospray ionization Key words: laser desorption, laser ionization, mass spec (Fenn et al., 1990) and matrix assisted la.5er desorption trometry, biomolecules, peptides, proteins, nucleotides, (Karas et al., 1989a) with m/z = 133,000 and m/z = nucleosides, saccharides, energy transfer, phase transi 250,000 high mass records , respectively. Th e latest de tion, disintegration, fragmentation, ion formation. velopments indicate capabilities for both techniques even *Address for correspondence: beyond these limits (Nohmi and Fenn, 1990; Williams A. Vertes and Nelson, 1990). Department of Chemistry, University of Antwerp On the other hand, one should not overemphasize (U.I.A.) , Universiteitsplein 1, B-2610 Wilrijk, Belgium the value of molecular weight determination by these Phone: (32)(3) 820.23.82, Fax : (32)(3) 820.23.76 methods . Several other, long establis hed techniques Electronic mail : [email protected] .be are available in the same or even in broader molecu lar weight range. Ultracentrifugation, light scattering, 317 A. Vertes and R. Gijbels gel permeation chromatography and gel electrophoresis existing ideas and models which try to explain laser de cover the molecular weight region from about 1,000 to sorption of large molecules. The concepts range from about 5,000,000 (Siggia, 1968). Th e cost and availability rapid heating (Beuhler et al. 1974) to mechanical ap of th e necessary instrum entation compares favorably to proaches (Williams, 1990), to restricted energy transfer the mass spectrometric equipment. When assessing the (Vertes, 1990) and to expansion cooling of the laser gen different molecular weight determination methods it is erated plume (Vertes et al., 1989b, Nelson et al. , 1989). necessary to keep in mind that mass spectrometry pro Las er indu ced thermal desorption vides directly mass to charge ratio data whereas the sep aration techniques measure particle mobilities and may The idea of laser indu ced thermal desorption incorporate systematic errors. (LITD) stems from early studies of the influ ence of laser An important aspect of the analytical value of the radiation on small molecul es adsorbed on solid surfa ces. mass spectrometric methods is their remarkably high If the substrate absorbs the laser radiation , it heats up sensitivities and low detection limits. Recent report s on a time-scale comparable to the lase r puls e leng th. The on electrospray ionization put sensitivities in the low resulting temp erature rise leads to the detachment of the pmol/ µL and detection limits in the low fmol region for adsorbed mol ecules. several 556 < m/z < 66,000 peptides (Van Berke! et al., It was realized soon that Q-switch ed laser pulses 1990). Both sensitivity and detection limits, however, ( durations are on th e ns time-scale) may lead to sub are highly compound dependent. The detection limit thermal velocity distribution s of the desorbed particl es increases with increasing molecular weight, because the (Wedler and Ruhmann, 1982). Furth er st udies suggested ion signal is distributed over more charged states (Loo that the desorption proce ss is adiabatic, espec ially in the et al., 1990). case of monol ayers (Simpson and Hardy , 1986). Similar , or even bette r results were obtained with The possibility to desorb cold molecul es from hot laser desorption . The sensitivity was in the low pmol/ µL, surfaces seemed very attractive and triggered st udi es on whereas the detection limits in the sub-pmol (Karas et larg er molecul es. Th e conflict of volatilization versus al., 1990) (10,000 < m/z < 100,000 peptides), or in the thermal !ability of these compounds was thought to be sub-fmol region (Hahn et al., 1987) (for porphyrin deriva overcome (van der Peyl et al., 1982a). Calculated sur tives). face temperatur es showed strong correlation with th e Naturally, these numb ers should be evaluated with generated ion curr ent s. To impl ement the met hod for the achievements of other techniqu es in mind . As an higher mass compound s, however , was apparently not example, we quote here some results on the analytical completely successful: m/z ~ 10,000 seemed to be an up performance of open tubular liquid chromatography. As per limit of ion produ ct ion by LITD (Ijam es and Wilkins , littl e as 4 fmol prot ein was hydrolysed and deriv at ized 1988). resulting in approximately 25 nL volume analyte solu Clearly , the evolution of substrat e surfac e tempera tion. Quantitativ e determination of 14 an1ino acid con ture plays a decisive role in th e course of events in LITD . st ituents was carried out with about 5 % error (Oates Tempo ral and spat ial temperature distributions, T(r,t) , and Jorgenson, 1990). Chromatographic and immunoa s in the sub strat e can be described by the heat conduction say analysis are serious competitors in terms of sens itiv equations: ity , with usually much lower cost of instrum ent at ion and consequently with better availability. Adv entur es to volatilize large mol ecules and de C(T) EJT(r, t) - divJ(r, t) + l (r , t), (1) tect minute quantiti es with the aid of lasers and mass at spectrometry will be the topic of this revi ew. Laser desorption mass spectro metry of nonvolatile organic molecules was surveyed by Shibanov (Shibanov 1986), J (r, t) = K (T )VT (r, t) (2) but vigorous development in the field since then has changed the landscape considerably. where I(r,t) is the source term describing the laser heat We will focus our attention on what the different ing and J(r,t) is th e heat flow. C(T) and K(T) are th e laser desorption and ionization methods have to offer in specific heat and the heat conductivity, respectively, gen excess of the capabilities of the more widespread analyt erally exhibiting considerable temperature depend ence. ical tools. Special emphasis will be given to the mass The source term can be expressed in terms of laser irra range and mass accuracy, to detection limits and sensi diance , I0 (r ,t), corrected for surface reflection: tivity, to the freedom from matrix interferences and to the availability of structural information. Two types of I(r, t) = alo(r, t)exp[-a z] (3) experiments will be distinguished in separate sections: laser induced thermal desorption and matrix assisted where the light propagates along the z axis perpendic laser desorption.

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