RAPID COMMUNICATIONS IN MASS SPECTROMETRY Rapid Commun. Mass Spectrom. 2007; 21: 1767–1775 Published online in Wiley InterScience (www.interscience.wiley.com) DOI: 10.1002/rcm.3011
Characterization of the chemical components on the surface of different solids with electrospray-assisted laser desorption ionization mass spectrometry Min-Zong Huang1, Hsiu-Jung Hsu1, Chen-I Wu1, Shu-Yao Lin1, Ya-Lin Ma1, Tian-Lu Cheng2,3 and Jentaie Shiea1,3* 1
Department of Chemistry, National Sun Yat-Sen University, Kaohsiung, Taiwan Department of Biomedical Science and Environmental Biology, Kaohsiung Medical University, Kaohsiung, Taiwan 3 National Sun Yat-Sen University-Kaohsiung Medical University Joint Center 2
Received 2 December 2006; Revised 25 March 2007; Accepted 26 March 2007
In this study we demonstrate that electrospray-assisted laser desorption ionization (ELDI) mass spectrometry (MS) can be used to rapidly characterize major chemical components on the surfaces of different solids under ambient conditions. The major chemical components in (a) dried milks with different fat contents, (b) different color-regions of a painting, (c) the thin coating on a compact disc, (d) drug tablets, and (e) porcine brain tissue were rapidly characterized as protonated molecules [MRH]R or sodiated molecules [MRNa]R by ELDI-MS with minimum sample pretreatment. The ionized ions of synthetic polymer and dye standards were detected directly from dried sample solutions using either positive or negative ion mode. Further structural information for the FD&C Red dye was obtained through tandem mass spectrometric (MS/MS) analysis using an ion trap mass analyzer attached to the ELDI source. Copyright # 2007 John Wiley & Sons, Ltd. The development of ambient ionization sources for characterizing organic and biological compounds directly from solid samples has been an important challenge for mass spectrometry (MS). Because such techniques would require essentially little or no sample pretreatment, their use in analytical procedures would be both time- and labor-saving, making it possible to apply them to high-throughput measurements. To date, the development of such ionization sources has focused mainly on impacting solid samples with protons, photons, or charged droplets for desorption/ ionization.1–5 Desorption electrospray ionization (DESI) is capable of desorbing and ionizing not only small organic but also large biological compounds directly from solid surfaces under ambient conditions.5–11 The technique is based on impinging the solid sample surface with a pneumatically assisted electrospray and then collecting the secondary ions generated through the interaction of charged microdroplets or gas-phase ions derived from the electrospray with molecules of the analyte present on the surface. The technique may still suffer, however, from a lack of high spatial resolution when imaging samples; in addition, the energy provided when impinging the sample surface with the charged droplets may not be sufficient to desorb the chemicals from a hard surface or biochemicals tightly bound within tissues. Recently, we reported a new desorption ionization method – electrospray-assisted laser desorption ionization (ELDI) – *Correspondence to: J. Shiea, Department of Chemistry, National Sun Yat-Sen University, Kaohsiung, 804 Taiwan. E-mail:
[email protected] Contract/grant sponsor: National Science Council, Taiwan, and Aim for the Top University Plan, Ministry of Education, Taiwan.
that combines some of the features of ESI and LD.12,13 This technique provides the advantages of allowing the direct, sensitive, and rapid characterization of protein standards in dried solutions under ambient conditions. The principle of ELDI is based on one of our previous designs – fused-droplet electrospray ionization (or two-step ESI).14–16 In a fuseddroplet ESI source, gaseous analyte molecules or neutral droplets containing the analyte molecules are conducted to the tip of an electrosprayer, where the analyte molecules are post-ionized through fusion or reaction with charged solvent droplets or protonated solvent species in the ESI plume. One of the advantages of using fused-droplet ESI for sample analysis is that the ionization and nebulization processes are separate events; this feature provides independent control over the conditions of the sample solution and the composition of the ESI solvent. By varying the methods of introducing the sample into a fused-droplet ESI source, unique applications have been demonstrated for liquid, gas, and solid sample analyses.17–21 Several investigators have noted previously that the number of neutral species desorbed by a laser pulse far exceeds the number of ions – and the desorbed neutral species are present for much longer time periods than are ions – after the laser pulse has been completed.22,23 Therefore, post-ionization of laser-desorbed neutrals remains an important research topic because this approach promises not only the detection of different analytes, but also increased sensitivity.24–29 In essence, the ELDI source is a modified
Copyright # 2007 John Wiley & Sons, Ltd.
1768 M.-Z. Huang et al.
fused-droplet ESI source that uses laser irradiation to produce the gaseous analyte molecules (or particles). The desorbed analyte molecules then join the electrospray plume, where they are post-ionized by the charged solvent species. Because ELDI can provide a large energy input to desorb the analyte molecules from the sample, this technique should be useful for the rapid characterization of chemicals directly from solids having either hard or soft surfaces under ambient conditions. In this paper, we report the use of ELDI-MS to characterize positively and negatively charged molecules directly from the surfaces of various solids, including (1) dried solutions of FD&C dyes, synthetic polymers, and milk with different fat contents; (2) active ingredients in drug tablets; and (3) different color regions of a painting, the thin film coated on a compact disc and porcine brain tissue. We also demonstrate that ELDI tandem mass spectrometry (MS/ MS) can be used for structural characterization – in this case for the FD&C Red dye molecule.
EXPERIMENTAL The standards, including three FD&C dyes, and synthetic polymers, and organic solvents (HPLC grade) were purchased from Sigma or Aldrich (Milwaukee, WI, USA) and used without further purification. The sample solution (ca. 10 mL of the standard solution or milk) was spread uniformly by pipetting it over a ca. 0.6-cm2 surface area (2 cm 0.3 cm) of a stainless-steel sample plate (5 cm in length 2 cm in width). The sample solution was then air-dried and subjected to ELDI-MS analysis. The painting and compact disc (CD) were
obtained from a local supermarket. The selected color region of a painting was cut and placed directly on the sample plate; the CD was cut into small pieces with a knife and the selected pieces were placed on the sample plate; the drug tablet was held on the sample plate using double-sided tape; all the samples were then analyzed by ELDI-MS without any further sample pretreatment. For analysis of the porcine brain tissue, the selected tissue area (ca. 10 mm 2 mm 1 mm height) was sliced off using a razor blade and placed on the sample plate prior to ELDI-MS analysis. The set-up of the ELDI instrument for this study is displayed in Fig. 1. The sample plate was positioned on a XYZ-stage in front of the sampling skimmer of an ion trap (Esquire 3000 plus; Bruker Daltonics, Billerica, MA, USA) or quadrupole time-of-flight (Q-TOF) (Bruker Daltonics BioTOF-q) mass analyzer. The selected sample area was then irradiated with a pulsed nitrogen laser (337 nm) operated at 10 Hz (controlled by a sweep function generator) using a pulsed energy of ca. 150 mJ and a pulse length of 4 ns. The laser beam (spot size: ca. 100 mm 150 mm) was focused through an objective lens. To prevent the analyte molecules on the dried sample spot being depleted by laser ablation or the surface of the porcine brain tissue from charring under laser irradiation, the sample plate was moved manually (ca. 0.2 mm/s) during data acquisition to ensure that fresh sample areas were probed. The mass spectra presented herein are averaged from ca. 200 laser shots collected from different positions. The laser-ablated molecules were post-ionized in the electrospray solvent plume. A methanol/water solution
Figure 1. (a) Photograph of the ELDI set-up. (b) Inset photograph displaying the relative positions of the sample plate (pre-applied with a droplet containing a pigment standard), capillary electrosprayer, and sampling skimmer of an ion trap mass analyzer. A: nitrogen laser cartridge (337 nm, 150 mJ); B: capillary connected to a syringe pump and a syringe filled with acidic MeOH solution (150 mL/h); C: ion trap mass analyzer; D: focusing lens; E: reflecting lens; F: sample plate attached on a three-way stage; G: three-way tee; H: sampling skimmer of the ion trap mass analyzer; I: sample plate possessing a stainless-steel surface (5 cm 2 cm); J: ESI capillary (150 mm i.d.); K: sample droplet. Copyright # 2007 John Wiley & Sons, Ltd.
Rapid Commun. Mass Spectrom. 2007; 21: 1767–1775 DOI: 10.1002/rcm
Characterization of chemical components on solid surfaces
TIC and observed no myoglobin ions in the mass spectra (Figs. 2(a) and 2(e)). When the laser power was switched on (indicated by the arrows in Fig. 2(a)), however, after a 2 s delay, the TIC increased abruptly by ca. 60-fold (ca. 8000 counts) and the ELDI mass spectrum displayed a complete multiply charged myoglobin ion series (Figs. 2(a) and 2(d)). Figures 2(b) and 2(c) display the change in the extracted ion current (EIC) of two myoglobin ions at m/z 1061 (þ16 charges) and m/z 1131 (þ15 charges), respectively, after laser irradiation. The changes in the TIC, EIC, and the mass spectra in Fig. 2 demonstrate unambiguously that the myoglobin molecules in the solid state were desorbed through nitrogen laser ablation and, subsequently, post-ionized in the electrospray plume. Because the ESI plume is aligned parallel to the sample plate and because we observed no myoglobin ions in the absence of laser irradiation, we believe that DESI-like ion formation processes do not occur during ELDI. Although it is not demonstrated in this study, the results of previous studies indicate that (1) reproducible protein ion signal can be generated in ELDI; (2) the detection limit of ELDI for protein analysis is ca. 107 M; and (3) the types of ions detected will be affected by the composition (or polarity) of the electrospray solution.12–14,30,31 Our success at obtaining protein ion signals from dried standard solutions and biological fluids in our previous studies suggests that large biomolecules in dried biological fluids can be rapidly characterized by ELDI-MS.13 To test the capability of ELDI-MS for the analysis of the biological fluids containing both small and large molecules, milk samples with different fat contents (whole and low-fat cow milk (
