Murray Group News – Page 19 – Murray Mass Spectrometry Group

Two-Laser Mid-Infrared and Ultraviolet Matrix-Assisted Laser Desorption/Ionization

Integrated ion signal for the NO loss fragment of 4-nitroaniline

M.W. Little, K.K. Murray, “Two-Laser Mid-Infrared and Ultraviolet Matrix-Assisted Laser Desorption/Ionization,” Int. J. Mass Spectrom. 261 (2007) 140–145. doi:10.1016/j.ijms.2006.08.010.

Abstract

Matrix-assisted laser desorption/ionization (MALDI) was performed using two-pulsed lasers with wavelengths in the infrared and ultraviolet regions. A 2.94 μm pulsed optical parametric oscillator laser system and a 337 nm pulsed nitrogen laser irradiated the same spot on the sample target. Sinapinic acid (SA), 2,5-dihydroxybenzoic acid (DHB), α-cyano-4-hydroxycinnamic acid (CCA), and 4-nitroaniline (NA) were used as matrices, and bovine insulin and cytochrome C were used as analytes. The laser energy was adjusted so that one-laser MALDI and LDI was at a minimum and the matrix and analyte ion signal was enhanced when the two lasers were fired together. Two-laser LDI was observed with SA, DHB, and NA matrices and two-laser MALDI was observed with SA and DHB. Plots of ion signal as a function of delay between the IR and UV lasers show two-laser signal from 0 ns up to a delay of 500 ns when the IR laser is fired before the UV laser. The results are interpreted in terms of IR laser heating of the target that leads to an enhancement in UV LDI and MALDI.

Wavelength dependence of soft infrared laser desorption and ionization

Inverse threshold fluence for insulin ion formation plotted as a function of wavelength

M.W. Little, J. Laboy, K.K. Murray, Wavelength dependence of soft infrared laser desorption and ionization, J. Phys. Chem. C. 111 (2007) 1412–1416. doi:10.1021/jp063154v.

Abstract

Protonated insulin molecules were formed by soft IR laser desorption ionization of a thin film of the protein on a silicon surface. Time-of-flight mass spectra were recorded at wavelengths between 2.8 and 3.6 µm and the efficiency of ionization was compared to the IR absorption of the protein thin film. Ionization efficiency was quantified by recording the minimum laser energy per unit area required to produce a detectable ion signal (threshold fluence). The ionization efficiency tracks the IR absorption spectrum of insulin between 2.6 and 3.8 µm in the region of OH, NH, and CH stretch absorption. The lowest threshold fluence and therefore the most efficient ionization was nearly coincident with the OH stretch absorption of insulin near 3.0 µm. An additional local maximum in ionization efficiency was observed at 3.4 µm, coincident with the CH stretch vibrational absorption. Comparison of the ionization efficiency with the IR absorption indicates that the protein and not the residual solvent is absorbing the laser energy. Scanning electron microscopy images of the bovine insulin thin films on silicon after laser irradiation show melting and indications of explosive boiling. Ionization occurs through the sacrifice of some of the protein molecules that absorb the laser energy and act as an intrinsic matrix.

Particle formation by infrared laser ablation of glycerol: implications for ion formation

Particle sized distribution weighted by count and mass from ablation of glycerol

S.N. Jackson, J.-K. Kim, J.L. Laboy, K.K. Murray, Particle formation by infrared laser ablation of glycerol: implications for ion formation, Rapid Commun. Mass Spectrom. 20 (2006) 1299–1304. doi:10.1002/rcm.2443.

Abstract

The quantity and size distribution of micrometer-sized particles ejected from thin films of glycerol were measured using light scattering particle sizing. Thin glycerol films were irradiated at atmospheric pressure with an infrared optical parametric oscillator at wavelengths between 2.95 and 3.1 µm. Particulate material resulting from the ablation was sampled directly into a particle-sizing instrument and particles with diameters greater than 500 nm were detected and sized by light scattering. The fluence threshold for particle formation was between 2000 and 3000 J/m² for all laser wavelengths. At threshold, fewer than 100 particles/cm³ were detected and this value increased to several thousand particles/cm³ at twice the threshold fluence. The average size of the coarse particles ranged from 900 nm to 1.6 µm at threshold and decreased by 10-20% at twice the threshold fluence. The coarse particle formation observations were compared with ion formation behavior in matrix-assisted laser desorption ionization and interpreted in terms of a photomechanical mechanism for material ablation and ion formation.

IR-MALDI-LDI combined with ion mobility orthogonal time-of-flight mass spectrometry

A.S. Woods, M.V. Ugarov, S.N. Jackson, T. Egan, H.-Y.J. Wang, K.K. Murray, J. A. Schultz, “IR-MALDI-LDI combined with ion mobility orthogonal time-of-flight mass spectrometry,” J. Proteome Res. 5 (2006) 1484–1487. doi:10.1021/pr060055l.

Abstract

Most MALDI instrumentation uses UV lasers. We have designed a MALDI−IM−oTOF−MS which employs both a Nd:YAG laser pumped optical parametric oscillator (OPOTEK, λ = 2.8−3.2 μm at 20 Hz) to perform IR−LDI or IR−MALDI and a Nd:YLF laser (Crystalaser, λ = 249 nm at 200 Hz) for the UV. Ion mobility (IM) gives a fast separation and analysis of biomolecules from complex mixtures in which ions of similar chemical type fall along well-defined “trend lines”. Our data shows that ion mobility allows multiply charged monomers and multimers to be resolved; thus, yielding pure spectra of the singly charged protein ion which are virtually devoid of chemical noise. In addition, we have demonstrated that IR−LDI produced similar results as IR−MALDI for the direct tissue analysis of phospholipids from rat brain.

Interfacing capillary gel microfluidic chips with infrared laser desorption mass spectrometry

Y. Xu, M.W. Little, K.K. Murray, “Interfacing capillary gel microfluidic chips with infrared laser desorption mass spectrometry,” J. Am. Soc. Mass Spectrom.17 (2006) 469–474. doi:10.1016/j.jasms.2005.12.003.

Abstract

Capillary gel microfluidic chip interfaced to laser desorption/ionization (LDI) mass spectrometry with a time-of-flight mass analyzer.

We report on the fabrication and performance of a gel microfluidic chip interfaced to laser desorption/ionization (LDI) mass spectrometry with a time-of-flight mass analyzer. The chip was fabricated from poly(methylmethacrylate) with a poly(dimethyl siloxane) cover. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis was performed in the channel of the microfluidic chip. After electrophoresis, the cover was removed and either the PDMS chip or the PMMA cover was mounted in a modified MALDI ion source for analysis. Ions were formed by irradiating the channel with 2.95 µm radiation from a pulsed optical parametric oscillator (OPO), which is coincident with IR absorption by N-H and O-H stretch of the gel components. No matrix was added. The microfluidic chip design allowed a decrease in the volume of material required for analysis over conventional gel slabs, thus enabling improvement in the detection limit to a pmol level, a three orders of magnitude improvement over previous studies in which desorption was achieved from an excised section of a conventional gel.

Matrix-assisted laser desorption/ionization mass spectrometry of collected bioaerosol particles

SEM of aerosol deposition on MALDI target

J.-K. Kim, S.N. Jackson, K.K. Murray, Matrix-assisted laser desorption/ionization mass spectrometry of collected bioaerosol particles, Rapid Commun. Mass Spectrom.19 (2005) 1725–1729. doi:10.1002/rcm.1982.

Abstract

A method was developed for collection and analysis of bioaerosols by matrix-assisted laser desorption/ionization (MALDI) time-of-flight mass spectrometry using a modified Andersen N6 bioaerosol collector. The overall goal of the study was to develop methods for obtaining mass spectra with minimal reagents and treatment steps for potential use in remote collection and analysis systems. Test bioaerosol particles were generated from a nebulized E. coli bacterial suspension and collected on MALDI targets placed in an Andersen N6 single-stage aerosol impactor. The bioaerosols were mixed with matrix either by deposition on a bare target with the matrix solution added later, or by deposition on a target pre-coated with matrix. The matrix compounds ?-cyano-4-hydroxycinnamic acid (CHCA) and sinapic acid (SA) were tested and the SA matrix was found to give the best results in number of peaks, resolution, and signal-to-noise ratio. Deposition of bioaerosol particles onto the matrix pre-coated target did not produce signal in the m/z region above 1000, but the signal could be recovered with the addition of a 1:1 (v/v) acetonitrile/water solvent. Addition of solvent by pipette to the pre-coated targets after particle deposition recovered signal comparable to the dried-droplet sample preparations, whereas solvent sprayed into the impactor recovered fewer peaks. Deposition on pre-coated targets with post-collection solvent addition was superior to deposition on bare target followed by post-collection addition of matrix solution.

Direct coupling of polymer-based microchip electrophoresis to online MALDI-MS using a rotating ball inlet

H. Musyimi, J. Guy, D.A. Narcisse, S.A. Soper, K.K. Murray, Direct coupling of polymer-based microchip electrophoresis to online MALDI-MS using a rotating ball inlet, Electrophoresis. 26 (2005) 4703–4710. doi:10.1002/elps.200500317.

Abstract

Rotating ball inlet with capillary electrophoresis microfluidic chip.

We report on the coupling of a polymer-based microfluidic chip to a MALDI-TOF MS using a rotating ball interface. The microfluidic chips were fabricated by micromilling a mold insert into a brass plate, which was then used for replicating polymer microparts via hot embossing. Assembly of the chip was accomplished by thermally annealing a cover slip to the embossed substrate to enclose the channels. The linear separation channel was 50 microm wide, 100 microm deep, and possessed an 8 cm effective length separation channel with a double-T injector (V(inj) = 10 nL). The exit of the separation channel was machined to allow direct contact deposition of effluent onto a specially constructed rotating ball inlet to the mass spectrometer. Matrix addition was accomplished in-line on the surface of the ball. The coupling utilized the ball as the cathode transfer electrode to transport sample into the vacuum for desorption with a 355 nm Nd:YAG laser and analyzed on a TOF mass spectrometer. The ball was cleaned online after every rotation. The ability to couple poly(methylmethacrylate) microchip electrophoresis devices for the separation of peptides and peptide fragments produced from a protein digest with subsequent online MALDI MS detection was demonstrated.

Matrix-free infrared soft laser desorption/ionization

D. Rousell, S.M. Dutta, M.W. Little, K.K. Murray, Matrix-free infrared soft laser desorption/ionization, J. Mass Spectrom.39 (2004) 1182–1189. doi:10.1002/jms.706.

Abstract

Infrared soft laser desorption/ionization was performed using a 2.94 µm Er:YAG laser and a commercial reflectron time‐of‐flight mass spectrometer. The instrument was modified so that a 337 nm nitrogen laser could be used concurrently with the IR laser to interrogate samples. Matrix‐assisted laser desorption/ionization (MALDI), laser desorption/ionization and desorption/ionization on silicon with UV and IR lasers were compared. Various target materials were tested for IR soft desorption ionization, including stainless steel, aluminum, copper, silicon, porous silicon and polyethylene. Silicon surfaces gave the best performance in terms of signal level and low‐mass interference. The internal energy resultant of the desorption/ionization was assessed using the easily fragmented vitamin B₁₂ molecule. IR ionization produced more analyte fragmentation than UV‐MALDI analysis. Fragmentation from matrix‐free IR desorption from silicon was comparable to that from IR‐MALDI. The results are interpreted as soft laser desorption and ionization resulting from the absorption of the IR laser energy by the analyte and associated solvent molecules.

IR Opolette and Er:YAG — IR Opolette OPO (front) and Er:YAG (back)

IR OPO and Er:YAG — IR Opolette OPO (left) and Er:YAG (right)

On-line single droplet deposition for MALDI mass spectrometry

X. Zhang, D.A. Narcisse, K.K. Murray, On-line single droplet deposition for MALDI mass spectrometry, J. Am. Soc. Mass Spectrom.15 (2004) 1471–1477. doi:10.1016/j.jasms.2004.06.016.

Abstract

A single droplet generator was coupled with a rotating ball inlet matrix-assisted laser desorption/ionization (MALDI) time of flight (TOF) mass spectrometer. Single droplets with 100 picoliter volume were ejected by a piezoelectric-actuated droplet generator and deposited onto a matrix-coated rotating stainless steel ball at atmospheric pressure. The single droplet deposit was transported to the vacuum side of the instrument where ionization was accomplished using a UV pulsed laser. Using this on-line interface, it was possible to obtain protonated molecule signal from as little as 10 fmol analyte.

Diagram of the single droplet deposition device: A, 19 mm diameter stainless steel ball; B, drive shaft; C, gasket; D, ISO 100 flange; E, ground grid; F, droplet generation electronics; G, syringe pump for matrix solution; H, syringe pump for solvent; I, capillary; J, translation stage for capillary; K, droplet generator; L, translation stage for droplet generator; M, cleaning system (Teflon holder and felt); N, peek plastic tube for waste; O, syringe pump for analyte solution.

On-line laser desorption/ionization mass spectrometry of matrix-coated aerosols

S.N. Jackson, S. Mishra, K.K. Murray, On-line laser desorption/ionization mass spectrometry of matrix-coated aerosols, Rapid Commun. Mass Spectrom.18 (2004) 2041–2045. doi:10.1002/rcm.1590.

Abstract

Matrix‐assisted laser desorption/ionization (MALDI) was used for the on‐line analysis of single particles. An aerosol was generated at atmospheric pressure and particles were introduced into a time‐of‐flight (TOF) mass spectrometer through a single‐stage differentially pumped capillary inlet. Prior to entering the mass spectrometer, a matrix was added to the particles using a heated saturator and condenser. A liquid matrix, 3‐nitrobenzyl alcohol (NBA), and a solid matrix, picolinic acid (PA), were used. Particles were ablated with a 351 nm excimer laser and the resulting ions were mass‐separated in a two‐stage reflectron TOF mass spectrometer. Aerosol particles containing the biomolecules erythromycin and gramicidin S were analyzed with and without the matrix addition step. The addition of NBA to the particles resulted in mass spectra that contained an intact molecular ion mass peak. In contrast, PA‐coated particles did not yield molecular ion peaks from matrix‐coated particles.

Aerosol MALDI instrument — Aerosol MALDI mass spectrometer schematic (top view)