Effect of the surface on charge reduction and desorption kinetics of soft landed peptide ions

Charge reduction and desorption kinetics of ions and neutral molecules produced by soft-landing of mass-selected singly and doubly protonated Gramicidin S (GS) on different surfaces was studied using time dependant in situ secondary ion mass spectrometry (SIMS) integrated in a specially designed Fourier transform ion cyclotron resonance mass spectrometer (FT-ICR MS) research instrument. Soft-landing targets utilized in this study included inert self-assembled monolayers (SAMs) of 1-dodecane thiol (HSAM) and its fluorinated analog (FSAM) on gold and hydrophilic carboxyl-terminated (COOH-SAM) and amine-terminated (NH₂-SAM) surfaces. We observed efficient neutralization of soft-landed ions on the COOH-SAM surface, partial retention of only one proton on the HSAM surface, and efficient retention of two protons on the FSAM surface. Slow desorption rates measured experimentally indicate fairly strong binding between peptide molecules and SAM surfaces with the binding energy of 20–25 kcal/mol.     

Reactive landing of peptide ions on self-assembled monolayer surfaces: an alternative approach for covalent immobilization of peptides on surfaces

Soft landing of mass-selected peptide ions onto reactive self-assembled monolayer surfaces (SAMs) was performed using a newly constructed ion deposition apparatus. SAM surfaces before and after soft landing were characterized ex situ using time-of-flight secondary-ion mass spectrometry (TOF-SIMS) and infrared reflection-absorption spectroscopy (IRRAS). We demonstrate that reactive landing (RL) results in efficient covalent linking of lysine-containing peptides onto the SAM of N-hydroxysuccinimidyl ester-terminated alkylthiol on gold (NHS-SAM). Systematic studies of the factors that affect the efficiency of RL revealed that the reaction takes place upon collision and is promoted by the kinetic energy of the ion. The efficiency of RL is maximized at ca. 40 eV collision energy. At high collision energies the RL efficiency decreases because of the competition with scattering of ions off the surface. The reaction yield is independent of the charge state of the projectile ions, suggesting that peptide ions undergo efficient neutralization upon collision. Chemical and physical properties of the SAM surface are also important factors that affect the outcome of RL. The presence of chemically reactive functional groups on the SAM surface significantly improves the reaction efficiency. RL of mass- and energy-selected peptide ions on surfaces provides a highly specific approach for covalent immobilization of biological molecules onto SAM surfaces.     

In situ SIMS analysis and reactions of surfaces prepared by soft landing of mass-selected cations and anions using an ion trap mass spectrometer

Mass-selected polyatomic cations and anions, produced by electrosonic spray ionization (ESSI), were deposited onto polycrystalline Au or fluorinated self-assembled monolayer (FSAM) surfaces by soft landing (SL), using a rectilinear ion trap (RIT) mass spectrometer. Protonated and deprotonated molecules, as well as intact cations and anions generated from such molecules as peptides, inorganic catalysts, and fluorescent dyes, were soft-landed onto the surfaces. Analysis of the modified surfaces was performed in situ by Cs⁺ secondary ion mass spectrometry (SIMS) using the same RIT mass analyzer to characterize the sputtered ions as that used to mass select the primary ions for SL. Soft-landing times as short as 30 s provided surfaces that yielded good quality SIMS spectra. Chemical reactions of the surfaces modified by SL were generated in an attached reaction chamber into which the surface was transferred under vacuum. For example, a surface on which protonated triethanolamine had been soft landed was silylated using vapor-phase chlorotrimethylsilane before being returned still under vacuum to the preparation chamber where SIMS analysis revealed the silyloxy functionalization. SL and vapor-phase reactions are complementary methods of surface modification and in situ surface analysis by SIMS is a simple way to characterize the products produced by either technique.     

In situ Raman spectroscopy of surfaces modified by ion soft landing

The design and characterization of a system for in situ Raman analysis of surfaces prepared by ion soft landing (SL) is described. The performance of the new high vacuum compatible, low cost, surface analysis capability is demonstrated with surface enhanced Raman spectroscopy (SERS) of surfaces prepared by soft landing of ions of crystal violet, Rhodamine 6G, methyl orange and copper phthalocyanine. Complementary in situ mass spectrometric information is recorded for the same surfaces using a previously implemented secondary ion mass spectrometer (SIMS). Imaging of the modified surfaces is achieved using 2D Raman imaging as demonstrated for the case of Rhodamine 6G soft landing. The combination of the powerful molecular characterization tools of SERS and SIMS in a single instrument fitted with in-vacuum sample transport capabilities, facilitates in situ analysis of surfaces prepared by ion SL. In particular, information is provided on the charge state of the soft landed species. In the case of crystal violet the SERS data suggest that the positively charged ions being landed retain their charge state on the surface under vacuum. By contrast, in the case of methyl orange which is landed as an anion, the SERS spectra suggest that the SL species has been neutralized.     

Studies on Fragmentation of Peptide by Nano-electrospray Ionization Fourier Transform Ion Cyclotron Resonance Multi-stage Tandem Mass Spectrometry

The sequence analysis of peptides was performed by nano-electrospray ionization Fourier transform ion cyclotron resonance tandem mass spectrometry(Nano-ESI-FT-ICR-MSn) and several peptides were chosen as examples. With the aid of the collision induced dissociation(CID), FT-ICR provides not only precise mass/charge ratio, but also structure information of the selected peptides. The fragment ions were identified according to the observed molecular weights and peptide sequence was determined successfully. So Nano-ESI-FT-ICR-MSn is a useful tool for identification of the amino acid sequence of peptides with high confidence. Besides, a pathway for the dehydration of y ions without amino acids containing carboxylic acid under sustained off-resonance irradiation collision-induced dissociation(SORI-CID) condition was proposed.     

Dynamics of energy transfer in peptide-surface collisions

Classical trajectory simulations are performed to study energy transfer in collisions of protonated triglycine (Gly)(3) and pentaglycine (Gly)(5) ions with n-hexyl thiolate self-assembled monolayer (SAM) and diamond [111] surfaces, for a collision energy E(i) in the range of 10-110 eV and a collision angle of 45 degrees. Energy transfer to the peptide ions' internal degrees of freedom is more efficient for collision with the diamond surface; i.e., 20% transfer to peptide vibration/rotation at E(i) = 30 eV. For collision with diamond, the majority of E(i) remains in peptide translation, while the majority of the energy transfer is to surface vibrations for collision with the softer SAM surface. The energy-transfer efficiencies are very similar for (Gly)(3) and (Gly)(5). Constraining various modes of (Gly)(3) shows that the peptide torsional modes absorb approximately 80% of the energy transfer to the peptide's internal modes. The energy-transfer efficiencies depend on E(i). These simulations are compared with recent experiments of peptide SID and simulations of energy transfer in Cr(CO)(6)(+) collisions with the SAM and diamond surfaces.     

Formation and electrochemical desorption of self‐assembled monolayers as studied by ToF‐SIMS

Time‐of‐flight secondary ion mass spectrometry (ToF‐SIMS) was used to study a number of processes involving thiol‐based self‐assembled monolayers (SAMs) on nontextured (polycrystalline) gold (Au) films deposited on Si wafers. ToF‐SIMS turned out to be a convenient and versatile semiqualitative technique which readily verified electrochemical desorption of a SAM and formation of another SAM on the same sample via reincubation with another thiol. The technique, allowing one to follow simultaneously more than one species on the surface, showed that any formation of a mixed SAM on surfaces which did not undergo electrolysis was negligible with the applied time scale (minutes). Copyright © 2010 John Wiley & Sons, Ltd.     

Fragmentation and reactivity in collisions of protonated diglycine with chemically modified perfluorinated alkylthiolate-self-assembled monolayer surfaces

Direct dynamics simulations are reported for quantum mechanical (QM)/molecular mechanical (MM) trajectories of N-protonated diglycine (gly(2)-H(+)) colliding with chemically modified perfluorinated octanethiolate self-assembled monolayer (SAM) surfaces. The RM1 semiempirical theory is used for the QM component of the trajectories. RM1 activation and reaction energies were compared with those determined from higher-level ab initio theories. Two chemical modifications are considered in which a head group (-COCl or -CHO) is substituted on the terminal carbon of a single chain of the SAM. These surfaces are designated as the COCl-SAM and CHO-SAM, respectively. Fragmentation, peptide reaction with the SAM, and covalent linkage of the peptide or its fragments with the SAM surface are observed. Peptide fragmentation via concerted CH(2)-CO bond breakage is the dominant pathway for both surfaces. HCl formation is the dominant species produced by reaction with the COCl-SAM, while for the CHO-SAM a concerted H-atom transfer from the CHO-SAM to the peptide combined with either a H-atom or radical transfer from the peptide to the surface to form singlet reaction products is the dominant pathway. A strong collision energy dependence is found for the probability of peptide fragmentation, its reactivity, and linkage with the SAM. Surface deposition, i.e., covalent linkage between the surface and the peptide, is compared to recent experimental observations of such bonding by Laskin and co-workers [Phys. Chem. Chem. Phys. 10, 1512 (2008)]. Qualitative differences in reactivity are seen between the COCl-SAM and CHO-SAM showing that chemical identity is important for surface reactivity. The probability of reactive surface deposition, which is most closely analogous to experimental observables, peaks at a value of around 20% for a collision energy of 50 eV.     

Assigning product ions from complex MS/MS spectra: The importance of mass uncertainty and resolving power

This study offers a unique insight into the mass accuracy and resolving power requirements in MS/MS analyses of complex product ion spectra. In the examples presented here, accurate mass assignments were often difficult because of multiple isobaric interferences and centroid mass shifts. The question then arose whether the resolving power of a medium-resolution quadrupole time-of flight (QqTOF) is sufficient or high-resolution Fourier-transform ion cyclotron resonance (FT-ICR) is required for unambiguous assignments of elemental compositions. For the comparison, two paralytic shellfish poisons (PSP), saxitoxin (STX) and neosaxitoxin (NEO), with molecular weights of 299 and 315 g x mol(-1), respectively, were chosen because of the high peak density in their MS/MS spectra. The assessment of QqTOF collision-induced dissociation spectra and FT-ICR infrared multiphoton dissociation spectra revealed that several intrinsic dissociation pathways leading to isobaric fragment ions could not be resolved with the QqTOF instrument and required FT-ICR to distinguish very close mass differences. The second major source of interferences was M + 1 species originating from coactivated 13C12Cc-1 ion contributions of the protonated molecules of the PSPs. The problem in QqTOF MS results from internal mass calibration when the MH+ ions of analyte and mass calibrant are activated at the same time in the collision or trapping cell. Although FT-ICR MS readily resolved these interfering species, the QqTOF did not provide resolving power >20,000 (full width at half maximum) required to separate most isobaric species. We were able to develop a semi-internal QqTOF calibration technique that activated only the isolated 12C isotope species of the protonated molecules, thus reducing the M + 1 interferences significantly. In terms of overall automated elemental formulas assignment, FT-ICR MS achieved the first formula hit for 100% of the product ions, whereas the QqTOF MS hit rate was only 56 and 65% for STX and NEO product ions, respectively. External mass calibration from commercial FT-ICR and QqTOF instruments gave similar results.     

Time of flight secondary ion mass spectrometric determination of molecular weight distributions of low polydispersity poly(dimethyl siloxane) with polyatomic primary ions

This work reports a comparison of oligomer and fragment ion intensities resulting from primary ion bombardment with several primary ion sources (Bi_n⁺, C₆₀⁺, and Cs⁺) at various energies in secondary ion mass spectrometry (SIMS). Although the use of polyatomic primary ions are of great interest due to increased secondary ion efficiency and yield, we demonstrate that monatomic primary ions result in increased oligomer ion yield for polymers prepared as submonolayer films on silver substrates. The enhancement of oligomer secondary ion yield with monatomic ions is evidence that monatomic primary ions have a shallower sampling depth than polyatomic ions, resulting from a collision cascade that is less energetic at the sample surface. The results are also consistent with a lower degree of fragmentation of the resultant secondary ions, which is observed when evaluating the fragmentation data and the spectral data.     

不同电荷态泛素蛋白离子的193 nm紫外光解离质谱

将193 nm激光与傅里叶变换离子回旋共振(FT-ICR)质谱仪结合, 研究了不同价态的泛素蛋白离子的紫外光解离质谱. 结果表明, 在光解离过程中向分析池内引入适量的碰撞气, 不仅能增加母体离子的裂解率, 也能提高碎片离子的捕获效率. 相对于碰撞辅助解离(CAD)中产生的b和y离子, 紫外光解离(UVPD)方法能够产生更为丰富的不同种类碎片离子. 其中, 对于+11价泛素离子, 蛋白质序列的覆盖率接近80%, 远高于对应的CAD实验结果. 与已往报道不同的是, 裂解覆盖率呈现出较强的电荷相关性. 因此, 如何进一步提高较低电荷态的蛋白离子的解离效率和序列覆盖率还需深入研究.     

High mass accuracy and high mass resolving power FT-ICR secondary ion mass spectrometry for biological tissue imaging

Biological tissue imaging by secondary ion mass spectrometry has seen rapid development with the commercial availability of polyatomic primary ion sources. Endogenous lipids and other small bio-molecules can now be routinely mapped on the sub-micrometer scale. Such experiments are typically performed on time-of-flight mass spectrometers for high sensitivity and high repetition rate imaging. However, such mass analyzers lack the mass resolving power to ensure separation of isobaric ions and the mass accuracy for elemental formula assignment based on exact mass measurement. We have recently reported a secondary ion mass spectrometer with the combination of a C₆₀ primary ion gun with a Fourier transform ion cyclotron resonance mass spectrometer (FT-ICR MS) for high mass resolving power, high mass measurement accuracy, and tandem mass spectrometry capabilities. In this work, high specificity and high sensitivity secondary ion FT-ICR MS was applied to chemical imaging of biological tissue. An entire rat brain tissue was measured with 150 μm spatial resolution (75 μm primary ion spot size) with mass resolving power (m/Δm _50%) of 67,500 (at m/z 750) and root-mean-square measurement accuracy less than two parts-per-million for intact phospholipids, small molecules and fragments. For the first time, ultra-high mass resolving power SIMS has been demonstrated, with m/Δm _50%?>?3,000,000. Higher spatial resolution capabilities of the platform were tested at a spatial resolution of 20 μm. The results represent order of magnitude improvements in mass resolving power and mass measurement accuracy for SIMS imaging and the promise of the platform for ultra-high mass resolving power and high spatial resolution imaging.

Shattering of Peptide ions on self-assembled monolayer surfaces

Time- and collision energy-resolved surface-induced dissociation (SID) of des-Arg(1)- and des-Arg(9)-bradykinin on a fluorinated self-assembled monolayer (SAM) surface was studied by use of a novel Fourier transform ion cyclotron resonance mass spectrometer (FT-ICR MS) specially equipped to perform SID experiments. Time-resolved fragmentation efficiency curves (TFECs) were modeled by an RRKM-based approach developed in our laboratory that utilizes a very flexible analytical expression for the internal energy deposition function capable of reproducing both single- and multiple-collision activation in the gas phase and excitation by collisions with a surface. Both experimental observations and modeling establish a very sharp transition in the dynamics of ion-surface interaction: the shattering transition. The experimental signature for this transition is the appearance of prompt (time-independent) fragmentation, which becomes dominant at high collision energies. Shattering opens a variety of dissociation pathways that are not accessible to slow collisional and thermal ion activation. This results in much better sequence coverage for the singly protonated peptides than dissociation patterns obtained with any of the slow activation methods. Modeling demonstrated that, for short reaction delays, dissociation of these peptides is solely determined by shattering. Internal energies required for shattering transition are approximately the same for des-Arg(1) and des-Arg(9)-bradykinin, resulting in the overlap of fragmentation efficiency curves obtained at short reaction delays. At longer delay times, parent ions depletion is mainly determined by a slow decay rate and fragmentation efficiency curves for des-Arg(1) and des-Arg(9)-bradykinin diverge. Dissociation thresholds of 1.17 and 1.09 eV and activation entropies of -22.2 and -23.3 cal/(mol K) were obtained for des-Arg(1) and des-Arg(9)-bradykinin from RRKM modeling of time-resolved data. Dissociation parameters for des-Arg(1)-bradykinin are in good agreement with parameters derived from thermal experiments. However, there is a significant discrepancy between the thermal data and dissociation parameters for des-Arg(9)-bradykinin obtained in this study. The difference is attributed to the differences in conformations that undergo thermal activation and activation by ion-surface collisions prior to dissociation.     

Energy deposition in desorption ionization

Deposition of internal energy accompanying the process of desorption ionization from a liquid matrix was assessed by comparing electrohydrodynamic ionization (EHMS) and fast atom bombardment (FABMS) mass spectra of glycerol solutions of a series of diquaternary ammonium salts. Observation of doubly-charged molecular parent ions in EHMS spectra confirmed the intrinsic stability of these ions in the gas phase. The more extensive fragmentation generally observed in the FABMS spectra suggests that the bombardment process increases the internal energy of sampled ions. However, the alternative explanation that instrumental (rather than sampling or ionization) differences prevent detection of fragments in EHMS cannot be ruled out. Similar considerations complicate comparison with published secondary ion (SIMS) spectra (obtained without liquid matrix), although the data indicate more extensive fragmentation with SIMS, consistent with other reports. Thus, deposition of internal energy accompanying ionization by these three desorption techniques may tentatively be ranked SIMS ⪢ FABMS ⪢ EHMS. Details of the EHMS spectra, including desolvation of solvated analyte cluster ions as a means of removing excess energy, were noted and discussed.     

Electrospray Ionization Fourier Transform Ion Cyclotron Resonance Mass Spectrometric Study on Sodium Azide Cluster Ions

Introduction Mass spectrometry (MS) has been an important technique for studying size-dependent cluster properties with unparalleled sensitivity, specificity, and versatil-ity.1,2 Especially Fourier transform ion cyclotron reso-nance (FT-ICR) MS offers the benefits of ultra-high mass resolving power, superior mass accuracy, and the ability of affording useful structure information with tandem MS. The introduction of electrospray ionization (ESI) as a gentle mode of ion generation for MS…     

High‐Resolution Surface Chemical Analysis of a Trifunctional Pattern Made by Sequential Colloidal Shadowing

We present a new method for creating surface chemical patterns where three chemistries can be periodically arranged at alternate positions on a single substrate without the use of top‐down approaches. High‐resolution chemical imaging by time‐of‐flight secondary ion mass spectrometry (ToF‐SIMS), with nanometer spatial resolution, is used to prove the success of the patterning and subsequent chemical modification steps. We use a combination of colloidal self‐assembly, plasma etching, self‐assembled monolayers (SAMs) and physical vapour deposition (PVD). The method utilizes a double colloid assembly process in which a first layer of close‐packed colloids is created, followed by plasma etching, coating with gold and deposition of a first SAM layer. A second particle layer is deposited on top of the first layer masking the interstitial spaces containing the first SAM. A second gold layer is deposited followed by a second SAM. After particle removal the surface consists of the pattern containing two different SAMs and a SiO₂ layer that can be readily functionalized with silanes. The possibility in the replacement of the two different thiols is investigated by X‐ray photoelectron spectroscopy (XPS) and it was found that no replacement is taking place. ToF‐SIMS imaging is used to show the periodicity of the chemical patterns by tracking unique fragment ions from the different surface regions. The patterning method is adaptable to create smaller or larger chemical patterns by appropriate choice of particle sizes. The patterns are useful for immobilizing biomolecules for cell studies or as multiplexed biosensors.     

SORI excitation: Collisional and radiative processes

Theoretical modeling of sustained off-resonance irradiation collision-induced dissociation (SORI-CID) experiments in Fourier transform ion cyclotron resonance (FT-ICR) mass spectrometry is described in the present paper. Manipulation of various analytical expressions yield the result that the average laboratory frame collision energy is equal to 2/3 of the maximum kinetic energy in SORI. Survival yields (the fraction of nondecomposed molecular ions) as a function of excitation time, collision energy, and source temperature have been considered: results of MassKinetics-type reaction kinetics modeling were compared with experimental results obtained by Guo et al. (Int. J. Mass Spectrom.2003, 225, 71-82). The results show that radiative cooling has a major influence in SORI-CID. They also suggest that collisional cooling is significant only at very low (less than 0.02 eV) center of mass collision energy; therefore it has a very small influence on the SORI process. Survival yield curves showed excellent agreement between experiments and calculations optimizing two parameters only (collisional energy-transfer efficiency and radiative cooling rate). Using leucine enkephalin as a model compound, the results indicate 0.128 +/- 0.021 energy deposition in a single collision and 7.5 +/- 0.5 s(-1) infrared cooling rate. We also present that these two physical parameters cannot be properly deconvoluted. This effect shows the importance of the parallel consideration of different physical processes.     

Energetics and dynamics of electron transfer and proton transfer in dissociation of metal(III)(salen)-peptide complexes in the gas phase

Time- and collision energy-resolved surface-induced dissociation (SID) of ternary complexes of Co(III)(salen)+, Fe(III)(salen)+, and Mn(III)(salen)+ with several angiotensin peptide analogues was studied using a Fourier transform ion cyclotron resonance mass spectrometer (FT-ICR MS) specially equipped to perform SID experiments. Time-resolved fragmentation efficiency curves (TFECs) were modeled using an RRKM-based approach developed in our laboratory. The approach utilizes a very flexible analytical expression for the internal energy deposition function that is capable of reproducing both single-collision and multiple-collision activation in the gas phase and excitation by collisions with a surface. The energetics and dynamics of competing dissociation pathways obtained from the modeling provides important insight on the competition between proton transfer, electron transfer, loss of neutral peptide ligand, and other processes that determine gas-phase fragmentation of these model systems. Similar fragmentation behavior was obtained for various Co(III)(salen)-peptide systems of different angiotensin analogues. In contrast, dissociation pathways and relative stabilities of the complexes changed dramatically when cobalt was replaced with trivalent iron or manganese. We demonstrate that the electron-transfer efficiency is correlated with redox properties of the metal(III)(salen) complexes (Co > Fe > Mn), while differences in the types of fragments formed from the complexes reflect differences in the modes of binding between the metal-salen complex and the peptide ligand. RRKM modeling of time- and collision-energy-resolved SID data suggests that the competition between proton transfer and electron transfer during dissociation of Co(III)(salen)-peptide complexes is mainly determined by differences in entropy effects while the energetics of these two pathways are very similar.     

Intermixed terpyridine-functionalized monolayers on gold: nonlinear relationship between terpyridyl density and metal ion coordination properties

Aiming at the functionalization of surfaces with terpyridine anchors for the coordinative deposition of additional layers, mixed self-assembled monolayers (SAMs) were prepared from binary solutions of 12-(2,2':6',2″-terpyridine-4'-yl)dodecane-1-thiol (TDT) and 1-decanethiol (DT). The SAMs and the order of the constituting molecules were analyzed by X-ray photoelectron spectroscopy (XPS), near-edge X-ray absorption fine structure spectroscopy (NEXAFS), and time-of-flight-secondary ion mass spectrometry (ToF-SIMS). The composition of the (TDT/DT)-SAMs and with it the surface density of terpyridyl groups correlates linearly with the relative concentrations of the two compounds in the solution used for depositing them. In marked contrast, the amount of terpyridine-coordinated Pd(II) ions significantly deviates from this trend with an optimum at a 1:3 ratio of TDT/DT. This indicates a major fraction of the terpyridines in TDT-rich SAMs not to be accessible for Pd(II) ion coordination. In agreement, NEXAFS spectroscopy reveals the alkyl backbones in TDT-rich SAMs not to be ordered, while they are preferentially upright oriented in the optimal 1:3-(TDT/DT)-SAMs. We interpret this in terms of terpyridine backfolding in TDT-rich SAMs, while they are located in accessible positions on top of the SAM in the 1:3-(TDT/DT)-SAM. While the alkyl backbones in the 1:3-(TDT/DT)-SAM are ordered, NEXAFS spectroscopy shows the terpyridyl groups not to have a preferential orientation in this SAM and thus retain enough flexibility to adjust to molecules that are deposited on top of the mixed SAM. In conclusion, the novel SAM does not undergo phase separation and consists predominantly of intermixed phases with adjustable surface density of quite flexible terpyridine anchor groups. The terpyridine-Pd(II) anchors are not only available for a future deposition of the next layer, but the metal ions also represent a sensitive probe for the accessibility of the terpyridyl groups.