Proteomics

In this work, we combined a newly developed matrix coating technique - matrix coating assisted by an electric field (MCAEF) and matrix-assisted laser desorption/ionization mass spectrometry (MALDI-MS) to enhance the imaging of peptides and proteins in tissue specimens of human prostate cancer. MCAEF increased the signal-to-noise ratios of the detected proteins by a factor of 2 to 5, and 232 signals were detected within the m/z 3500-37500 mass range on a time-of-flight mass spectrometer and with the sinapinic acid MALDI matrix. Among these species, three proteins (S100-A9, S100-A10, and S100-A12) were only observed in the cancerous cell region and 14 proteins, including a fragment of mitogen-activated protein kinase/extracellular signal-regulated kinase kinase kinase 2, a fragment of cAMP-regulated phosphoprotein 19, 3 apolipoproteins (C-I, A-I, and A-II), 2 S100 proteins (A6 and A8), β-microseminoprotein, tumor protein D52, α-1-acid glycoprotein 1, heat shock protein β-1, prostate-specific antigen, and 2 unidentified large peptides at m/z 5002.2 and 6704.2, showed significantly differential distributions at the p < 0.05 (t-test) level between the cancerous and the noncancerous regions of the tissue. Among these 17 species, the distributions of apolipoprotein C-I, S100-A6, and S100-A8 were verified by immunohistological staining. In summary, this study resulted in the imaging of the largest group of proteins in prostate cancer tissues by MALDI-MS reported thus far, and is the first to show a correlation between S100 proteins and prostate cancer in a MS imaging study. The successful imaging of the three proteins only found in the cancerous tissues, as well as those showing differential expressions demonstrated the potential of MCAEF-MALDI/MS for the in situ detection of potential cancer biomarkers. Copyright © 2015 John Wiley & Sons, Ltd.

LC coupled to ESI, MALDI and ICP MS – A multiple hyphenation for metalloproteomic studies

Pulsed laser ablation synthesis of fresh Te nanoparticles for matrix assisted laser desorption/ionization mass spectrometry (MALDI-MS) applications

Surface functionalization is considered to be the foundation for developing nanomaterial applications in matrix-assisted laser desorption/ionization mass spectrometry (MALDI-MS) analyses. However, the surface properties of nanostructures can influence their interaction with the analyte and consequently the mass data. In the present study, functionalized nanoparticles (NPs) were used for MALDI-MS and laser desorption/ionization mass spectrometry (LDI-MS) experiments in order to evaluate the effect of the surface properties of NPs on tailoring the intensity of mass signals. Regarding the LDI-MS analyses, the surface of superparamagnetic iron oxide nanoparticles (SPIONs) was coated with nitrosonium tetrafluoroborate, citric acid, nitrodopamine, and gallic acid. Additionally, the SPIONs were applied as a matrix to analyze three small peptides. In the MALDI-MS analyses, silica NPs were selected as co-matrix and functionalized with cysteine, sulfobetaine, and amine alkoxysilanes. Then, the silica NPs were utilized as additives in the MALDI-MS samples of four proteins in a mass range between ~2000 and 60,000 Da. The results of LDI-MS analyses demonstrated more than one order enhancement in the signal intensity of analytes based on the amount of electrostatic interaction and laser energy absorption by the surface ligands. However, those of MALDI-MS experiments indicated a significant signal improvement when achieving the colloidal stability of silica NPs in the matrix solution. Based on the results, the surface properties of NPs affected the (MA)LDI-MS analyses indispensably. Finally, the functionalization of SPIONs represented a new model for the future development of NPs with both affinity and enhanced ionization abilities in mass spectrometry.

To open up new ways for matrix-assisted laser desorption/ionization mass spectrometry (MALDI-MS)-based patient screening, blood serum is the most preferred specimen because of its richness in patho-physiological information and due to ease of collection. To overcome deleterious freeze/thaw cycles and to reduce high costs for shipping and storage, we sought to develop a procedure which enables MALDI-MS protein profiling of blood serum proteins without the need for serum freezing. Blood sera from patients/donors were divided into portions which after pre-incubation were fast frozen. Thawed aliquots were deposited on filter paper discs and air-dried at room temperature. Intact serum proteins were eluted with acid-labile detergent-containing solutions and were desalted by employing a reversed-phase bead system. Purified protein solutions were screened by MALDI-MS using standardized instrument settings. MALDI mass spectra from protein solutions which were eluted from filter paper discs and desalted showed on average 25 strong ion signals (mass range m/z 6000 to 10,000) from intact serum proteins (apolipoproteins, complement proteins, transthyretin and hemoglobin) and from proteolytic processing products. Semi-quantitative analysis of three ion pairs: m/z 6433 and 6631, m/z 8205 and 8916, as well as m/z 9275 and 9422, indicated that the mass spectra from either pre-incubated fast-frozen serum or pre-incubated dried serum spot eluted serum contained the same information on protein composition. A workflow that avoids the conventional cold-chain and yet enables the investigation of intact serum proteins and/or serum proteolysis products by MALDI-MS profiling was developed. The presented protocol tremendously broadens the clinical application of MALDI-MS and simultaneously allows a reduction in the costs for storage and shipping of serum samples. This will pave the way for clinical screening of patients also in areas with limited access to health care systems, and/or specialized laboratories.

While surfactants are commonly used in preparing protein samples, their presence in a protein sample can potentially affect the enzymatic digestion process and the subsequent analysis of the resulting peptides by mass spectrometry. The extent of the tolerance of matrix-assisted laser desorption/ionization mass spectrometry (MALDI-MS) to surfactant interference in peptide analysis is very much dependent on the matrix/sample preparation method. In this work the effects of four commonly used surfactants, namely n-octyl glucoside (OG), Triton X-100 (TX-100), 3-[(3-cholamidopropyl)dimethylammonio]-1-propanesulfonate (CHAPS) and sodium dodecyl sulfate (SDS), for biological sample preparation on trypsin digestion and MALDI-MS of the resulting digest are examined in detail within the context of using a two-layer method for MALDI matrix/sample preparation. Non-ionic and mild surfactants, such as OG, TX-100 or CHAPS, are found to have no significant effect on trypsin digestion with surfactant concentrations up to 1%. However, TX-100 and CHAPS interfere with the subsequent peptide analysis by MALDI-MS and should be removed prior to peptide analysis. OG is an MS-friendly surfactant and no effect is observed for MALDI peptide analysis. The effect of SDS on trypsin digestion in terms of the number of peptides generated and the overall protein sequence coverage by these peptides is found to be protein dependent. The use of SDS to solubilize hydrophobic membrane proteins, followed by trypsin digestion in the presence of 0.1% SDS, results in a peptide mixture that can be analyzed directly by MALDI-MS. These peptides are shown to provide better sequence coverage compared with those obtained without the use of SDS in the case of bacteriorhodopsin, a very hydrophobic transmembrane protein. This work illustrates that MALDI-MS with the two-layer sample preparation method can be used for direct analysis of protein digests with no or minimum sample cleanup after proteins are digested in a solution containing surfactants.

The development of simplified procedures for isolating high-mass alkanes present in crude oils is described. The new procedures, which bypass the sample recovery step with hot toluene in the conventional alkane-isolation procedure, also provide an effective sample preparation route, prior to analysis by matrix-assisted laser desorption/ionisation mass spectrometry (MALDI-MS). Urea-alkane adducts are formed by mixing sample and urea solutions on chromatographic paper or silica-coated plates. Unreacted hydrocarbons are removed by developing the plates with chloroform. In a second development with water, adducts are broken up in situ and the liberated urea removed, leaving bands of isolated alkanes behind. For MALDI-MS, strips of paper/plates, carrying the isolated alkanes, are fixed on metal target plates. The samples are treated with matrix (AgNO(3)) and analysed by MALDI-MS. The observed signal represents silver ion adducts of the isolated alkanes. Silver appears to work without much fragmentation and to generate whole silver adduct ions. Much improved MALDI-MS detection sensitivity and a wider range of masses was observed when samples were ablated from paper/plate surfaces, than by ablation from bulk samples spread over a smooth surface--the conventional method. Chromatographic paper gave better resolution and a broader range of masses than silica-coated plates. The analytical sequences have been confirmed using standard alkanes (C(20)-C(60)) and Polywax. The proposed procedures enhanced the sensitivity and detection range of the MS analysis. The method was useful in detecting n-alkanes to m/z 1500 (C(100)) and required relatively small quantities of sample and reagents. It provides a promising qualitative analysis route for the rapid isolation and reliable determination of alkanes in crude oils.

Herein, we re-explored α-cyano-4-hydroxycinnamic acid (CHCA) as a reactive matrix for selective and sensitive analysis of glutathione (GSH) by matrix-assisted laser desorption/ionization mass spectrometry (MALDI-MS). CHCA efficiently reacted with GSH, and the resulting CHCA-GSH conjugate was readily detected by MALDI-MS without interferences. The detection limit of the CHCA-GSH conjugate decreased to 200 pmol μL-1, which was 2 orders of magnitude lower than that of pure GSH.Forapplication, CHCA was successfully applied for the detection of GSH, present in HepG2 cell lysates. The results demonstrated detection advantages of simple, high-throughput, and selective and screening of GSH in biological samples by MALDI-MS.

Reaction of zinc dithiophosphate with dimethylsulfoxide was used to analyze zinc dialkyldithophosphates and diaryldithiophosphates lubricating oil additives by matrix assisted laser desorption ionization mass spectrometry (MALDIMS) analysis. The MALDI-MS analysis was carried out by mixing the [Zn(DTP)(DMSO)[subscript 2]][superscript +] with various type of matrix such as 2,5-dihydroxybenzoic acid, dithranol and alpha-cyano-4-hydroxycinnamic acid. The mixture of [Zn(DTP)(DMSO)2]+and matrix was applied to the multiprobe and bombarded with laser to genarate ionic analyte. The following MALDI-MS conditions were found to provided the good MALDI-MS spectrum of [Zn(DTP)(DMSO)[subscript 2]][superscript +] : dithranol as a matrix with tetrahydrofuran as solvent, the ratio of [Zn(DTP)(DMSO)[subscript 2]][superscript +] : matrix was 1 : 75 by volume and laser power was 187 [micro]J. The MALDI-MS spectrum of zinc dialkyldithiophosphates showed the peaks between m/z 517 and 615 and zinc diaryldithiophosphates showed the peaks between m/z 697 and 795. The MALDI-MS analysis of [Zn(DTP)(DMSO)2][superscript +] provided information on the carbon number of the two alkyl chains (R1+R2) which is an individual fingerprint of zinc dithiophosphates for any given commercial lubricating oils. Therefore, the developed MALDI-MS was used to analyze zinc dithiophosphates in commercial lubricating oils.

A simple method using matrix‐assisted laser desorption ionization mass spectrometry (MALDI‐MS) is described for the characterization of acylcarnitines in standard solutions and urinary samples from controls and patients with β‐oxidation deficiency disorders. This mass spectrometric approach is compared with other techniques currently utilized for studying acylcarnitines: fast atom bombardment mass spectrometry (FABMS), gas chromatography/chemical ionization mass spectrometry (GC/CIMS) and electrospray ionization mass spectrometry (ESI‐MS). MALDI‐MS was found to compare favourably with the recently described ESI‐MS analysis of acylcarnitines. Both ESI‐MS and MALDI‐MS share the following advantages: (i) non‐requirement of a derivatization step; (ii) easy detection of acylcarnitine esters in standard preparations and urinary samples from human subjects; (iii) detection of fatty acid oxidation disorders from the urinary acylcarnitine profiles in patients suffering from medium‐chain acyl‐CoA dehydrogenase deficiency (MCADD), multiple acyl‐CoA dehydrogenase deficiency (MADD) and mitochondrial carnitine palmitoyltransferase type II (CPT II) deficiency; (iv) sensitivity comparable to FABMS and GC/CIMS, the two reference methods most widely utilized for routine detection of acylcarnitines; and (v) ability (like FABMS but in contrast with GC/CIMS) to detect long‐chain acylcarnitines, suggesting that these esters might have been previously underestimated in body fluids from patients with β‐oxidation deficiency disorders. MALDI‐MS, like ESI‐MS, provides a fast and sensitive method for determining carnitine and its esters, with the potential for routine application in neonatal screening for inherited metabolic disorders involving acylcarnitine metabolism.

Matrix-assisted laser desorption/ionization mass spectrometry (MALDI-MS) has been a major approach for glycan analysis. However, the preferential cleavage of the sialic acid moiety by in- and post-source decay influences the determination of sialylated glycans by MALDI-MS. Many chemical derivatization methods were introduced to stabilize the sialylated glycan during MALDI-MS. Among current derivatization methods, methylamidation is a promising means for simultaneous analysis of natural sialylated glycans regardless of their sialic acid linkage types. Here, a novel derivatization method was developed, in which proteins were conjugated on the solid-phase support in order to stabilize the sialic acids by methylamidation and to reduce sample loss and contamination during the derivatization process. This derivatization strategy was used to investigate N-glycans from fetuin, a glycoprotein containing different types of complex N-glycans. The developed method was also applied to the N-glycan profiling of human serum from patients and healthy volunteers. Results were consistent with N-glycan profiling by HPLC-fluorescence detection. This new method provided a sensitive, simple, and robust approach for profiling glycan structures of complex samples.

Liquid chromatography coupled to mass spectrometry (LC-MS) is an excellent technique to determine trace levels of polar and thermolabile pesticides and their degradation products in complex matrices. LC-MS can be equipped with several mass analyzers, each of which provides unique features capable to identify, quantify, and resolve ambiguities by selecting appropriate ionization and acquisition parameters. We discuss in this review the use of LC coupled to (quadrupole) time-of-flight mass spectrometry (LC-(Q)ToF-MS) to determine the presence of target and non-target pesticides in water and food. This technique is characterized by operating at a resolving power of 10,000 or more. Therefore, it gives accurate masses for both parent and fragment ions and enables the measurement of the elemental formula of a compound achieving compound identification. In addition, the combination of quadrupole-ToF permits tandem mass spectrometry, provides more structural information, and enhances selectivity. The purpose of this article is to provide an overview on the state of art and applicability of liquid chromatography time-of-flight mass spectrometry (LC-ToF-MS), and liquid chromatography quadrupole time-of-flight mass spectrometry (LC-QToF-MS) for the analysis of pesticides in environmental matrices and food. The performance of such techniques is depicted in terms of accurate mass measurement, fragmentation, and selectivity. The final section is devoted to describing the applicability of LC-(Q)ToF-MS to routine analysis of pesticides in food matrices, indicating those operational conditions and criteria used to screen, quantify, and identify target and "suspected" pesticides and their degradation products in water, fruits, and vegetables. The potential and future trends as well as limitations of LC-(Q)ToF-MS for pesticide monitoring are highlighted.

Introduction Book Editor(s):Charles L. Brooks III, Charles L. Brooks III Department of Chemistry, Carnegie-Mellon University, Pittsburgh, PennsylvaniaSearch for more papers by this authorMartin Karplus, Martin Karplus Department of Chemistry, Harvard University, Cambridge, MassachusettsSearch for more papers by this authorB. Montgomery Pettitt, B. Montgomery Pettitt Department of Chemistry, University of Houston, Houston, TexasSearch for more papers by this author First published: 01 January 1988 https://doi.org/10.1002/9780470141205.ch1Citations: 136Book Series:Advances in Chemical Physics AboutPDFPDF ToolsRequest permissionExport citationAdd to favoritesTrack citation ShareShareShare a linkShare onFacebookTwitterLinked InRedditWechat Citing Literature Advances in Chemical Physics: Proteins: A Theoretical Perspective of Dynamics, Structure, and Thermodynamics, Volume 71 RelatedInformation

This invited Team Profile was created by Prof. Judy I. Wu, Department of Chemistry, University of Houston (USA) and Prof. Michael M. Haley, Department of Chemistry & Biochemistry and the Materials Science Institute, University of Oregon, Eugene (USA). The X-ray structure of tetra-t-butyl-s-indacene was reported in 1986 to possess a symmetric, delocalized D2h geometry, which went against conventional wisdom that this antiaromatic should exhibit a bond-localized C2h structure. Computational studies over the ensuing 35+ years yielded conflicting outcomes as to the preferred geometry. A computational reexamination of this structure by our team found experimental proton chemical shifts matched best with a computed C2h symmetry, confirming a bond-localized geometry. "Tetra-tert-butyl-s-indacene is a Bond-Localized C2h Structure and a Challenge for Computational Chemistry", L. J. Karas, S. Jalife, R. V. Viesser, J. V. Soares, M. M. Haley, J. I. Wu, Angew. Chem. Int. Ed. 2023, 62, e202307379.

Tissue imaging using matrix assisted laser desorption/ionization mass spectrometry (MALDI-MS) is a well-established technique that, in recent years, has seen wider adoption and novel application. Applications such imaging mass spectrometry (IMS) and biotyping are beginning to gain greater exposure and use; however, with limitations in optimization methods, producing the best result often relies on the ability to customize the physical characteristics of the instrumentation, a task that is challenging for most mass spectrometry laboratories. With this in mind, we have described the effect of making simple adjustments to the laser optics at the final collimating lens area, to adjust the laser beam size and shape in order to allow greater customization of the instrument for improving techniques such as IMS. We have therefore been able to demonstrate that improvements can be made without requiring the help of an electrical engineer or external funding in a way that only costs a small amount of time. Graphical Abstract ᅟ.