Matrix-assisted laser desorption ionization mass spectrometry for identification of shrimp

Matrix-assisted laser desorption ionization (MALDI) time of flight mass spectrometry was used to identify shrimp at the species level using commercial mass spectral fingerprint matching software (Bruker Biotyper). In the first step, a mass spectrum reference database was constructed from the analysis of six commercially important shrimp species: Litopenaeus setiferus, Farfantepenaeus aztecus, Sicyonia brevirostris, Pleoticus robustus, Pandalopsis dispar and Pandalus platyceros. This step required a desalting procedure for optimum performance. In the second step, the reference database was tested using 74 unknown shrimp samples from these six species. Correct identification was achieved for 72 of 74 samples (97%): 72 samples were identified at the species level and 2 samples were identified at the genus level using the manufacturer's log score specifications. The MALDI fingerprinting method for the identification of shrimp species was found to be reproducible and accurate with rapid analysis.     

Mass spectrometric analysis of low molecular mass polyesters by laser desorption/ionization on porous silicon

A low molecular mass polyester was analyzed by desorption/ionization on porous silicon (DIOS) mass spectrometry. The results were compared with those of matrix-assisted laser desorption ionization (MALDI) mass spectrometry using matrixes of alpha-cyano-4-hydroxycinnamic acid (CHCA) and 10,15,20-tetrakis(pentafluorophenyl)porphyrin (F20TPP). The CHCA matrix was not suitable for characterization of low molecular mass components of the polyester because the matrix-related ions interfered with the component ions. On the other hand, the F20TPP matrix showed no interference because no matrix-related ions appeared below m/z 822. However, the solvent selection for determining optimal conditions of sample preparation was limited, because F20TPP does not dissolve readily in any of the available organic solvents. In the DIOS spectra, the polymer ions were observed at high sensitivity without a contaminating ion. No matrix is needed for DIOS spectra of low molecular mass polyesters, facilitating sample preparation and selectivity of a precursor ion in post-source decay measurements.     

Study of synthetic aliphatic copolyamides by time-of-flight matrix assisted laser desorption/ionization mass spectrometry

Synthetic copolyamides based on aliphatic diamines (1,3-propanediamine and 1,4-butanediamine) and dichlorides of aliphatic carboxylic acids (adipic and sebacic acid dichlorides) were investigated using time-of-flight matrix assisted laser desorption/ionization mass spectrometry. Their mass spectra showed peaks for cationized (Na⁺ and K⁺) and protonated (less intense peaks) oligomers with NH₂-NH₂, NH₂-COOH, or COOH-COOH end groups. No cyclic oligomers were detected in the samples. The compositions of oligomers were determined, and the relative reactivities of homologous comonomers in polycondensation were estimated. Published in Russian in Izvestiya Akademii Nauk. Seriya Khimicheskaya, No. 7, pp. 1320–1324, July, 2007.     

1,8-Bis(dimethylamino)naphthalene/9-aminoacridine: A new binary matrix for lipid fingerprinting of intact bacteria by matrix assisted laser desorption ionization mass spectrometry

The effectiveness of a novel binary matrix composed of 1,8-bis(dimethylamino)naphthalene (DMAN; proton sponge) and 9-aminoacridine (9AA) for the direct lipid analysis of whole bacterial cells by matrix assisted laser desorption ionization mass spectrometry (MALDI MS) is demonstrated. Deprotonated analyte signals nearly free of matrix-related ions were observed in negative ion mode. The effect of the most important factors (laser energy, pulse voltage, DMAN/9AA ratio, analyte/matrix ratio) was investigated using a Box–Behnken response surface design followed by multi-response optimization in order to simultaneously maximize signal-to-noise (S/N) ratio and resolution. The chemical surface composition of single or mixed matrices was explored by X-ray photoelectron spectroscopy (XPS). Moreover, XPS imaging was used to map the spatial distribution of a model phospholipid in single or binary matrices.     

Effective detection of peptides containing cysteine sulfonic acid using matrix-assisted laser desorption/ionization and laser desorption/ionization on porous silicon mass spectrometry

Cysteine sulfonic acid-containing peptides, being typical acidic peptides, exhibit low response in matrix-assisted laser desorption/ionization (MALDI) mass spectrometry. In this study, matrix conditions and the effect of diammonium hydrogencitrate (DAHC) as additive were investigated for ionization of cysteine sulfonic acid-containing peptides in MALDI. A matrix-free ionization method, desorption/ionization on porous silicon (DIOS), was also utilized to evaluate the effect of DAHC. When equimolar three-component mixtures of peptides carrying free cysteine, cysteine sulfonic acid, and carbamidomethyl cysteine were measured by MALDI using a common matrix, alpha-cyano-4-hydroxycinnamic acid (CHCA), no signal corresponding to cysteine sulfonic acid-containing peptide could be observed in the mass spectrum. However, by addition of DAHC to CHCA, the peaks of cysteine sulfonic acid-containing peptides were successfully observed, as well as when using 2,4,6-trihydroxyacetophenone (THAP) and 2,6-dihydroxyacetophenone with DAHC. In the DIOS mass spectra of these analytes, the use of DAHC also enhanced the peak intensity of the cysteine sulfonic acid-containing peptides. On the basis of studies with these model peptides, tryptic digests of oxidized peroxiredoxin 6 were examined as a complex peptide mixture by MALDI and DIOS. In MALDI, the peaks of cysteine sulfonic acid-containing peptides were observed when using THAP/DAHC as the matrix, but this was not so with CHCA. In DIOS, the signal from cysteine sulfonic acid-containing peptides was suppressed; however, the use of DAHC significantly enhanced the signal intensity with an increase in the number of observed peptides and increased signal-to-noise ratio in the DIOS spectra. The results show that DAHC in the matrix or on the DIOS chip decreases discrimination and suppression effects in addition to suppressing alkali-adduct ions, which leads to a beneficial effect on protonation of peptides containing cysteine sulfonic acid.     

基质辅助激光解吸电离飞行时间质谱研究2: 四硫 富瓦烯化合物

采用基质辅助激光解吸电离飞行时间质谱(MALDI-TOF-MS),对四硫富瓦烯化合物进行质谱表征。在所用的实验条件下,样品很容易解吸电离生成单电荷分子离子,得到单同位素分辨的质谱图。26种实际样品的质谱分析结果表明;MALDI-TOF-MS可以比其它质谱方法更有效、更方便地用于此类化合物的质谱分析,解决了此类化合物不易进行质谱鉴定的难题。     

人参皂苷的基质辅助激光解吸质谱研究

利用MALDI-TOFMS测定了八种人参皂苷的分子量, 并分析了西洋参总皂苷的组成。同时, 进行了灵敏度实验, 并探讨了基质及碱金属离子的影响, 证明该方法灵敏度高, 重复性好, 结果准确。是测定极性小分子分子量的有效方法。     

Mesoporous tungsten titanate as matrix for matrix-assisted laser desorption/ionization time-of-flight mass spectrometry analysis of biomolecules

In this paper, mesoporous tungsten titanate (WTiO) with different nano-pore structures was utilized as matrix for the analysis of short peptides by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOFMS). Effect of characteristic features of mesoporous matrices on laser desorption/ionization process was investigated. Experiments showed that the ordered two-dimensional and three-dimensional mesoporous matrices were superior in performance to the non-ordered WTiO matrix. The dramatic enhancement of signal sensitivity by the ordered mesoporous matrices can be reasonably attributed to the ordered structure, which facilitated the understanding on structure-function relationship in mesoporous cavity for laser desorption process of adsorbed biomolecules. With the ordered mesoporous matrix, the short peptides are successfully detected. The presence of trace alkali metal salt effectively increased the analyte ion yields and the MALDI-TOFMS using the inorganic mesoporous matrices displayed a high salt tolerance. The developed technique also showed a satisfactory performance in peptide-mapping and amino-acid sequencing analysis.     

Capillary electrophoretic separation and fractionation of hydrophobic peptides onto a pre-structured matrix assisted laser desorption/ ionization target for mass spectrometric analysis

A CE separation of hydrophobic peptides followed by fractionation onto a prestructured MALDI target and off-line MS analysis was performed. An improved and partially automated manufacturing procedure of the previously described MALDI target is presented. This target is structurally coated with silicone and especially developed for hydrophobic peptides and proteins. Here, the target plate was designed specifically for the CE fraction collection. Different solvents were evaluated to meet the requirements of peptide solubility and compatibility to both the CE and MALDI methods and to the fractionation procedure. CE-MALDI-MS analysis of nine highly hydrophobic peptides from cyanogen bromide-digested bacteriorhodopsin is demonstrated.     

Automated normal phase nano high performance liquid chromatography/matrix assisted laser desorption/ionization mass spectrometry for analysis of neutral and acidic glycosphingolipids

The coupling of nano high-performance liquid chromatography (nanoHPLC) with matrix-assisted laser desorption/ionization (MALDI) mass spectrometry (MS) via an automatic spotting roboter was developed and adapted for the first time for the analysis of complex mixtures of glycosphingolipids (GSLs). The 2,5-dihydroxybenzoic acid and 6-azo-2-thiothymine matrix systems were adjusted to concurrently meet the requirements for reproducible and homogeneous crystal formation with the liquid chromatography (LC) eluent under the variable LC solvent composition over the course gradient and high ionization efficiency of the GSL species, without the need for recrystallization. Precise adjustment of the automatic spotting parameters in terms of matrix flow rate, on-tip collection time of the matrix/LC eluent solution and the matrix spotting mode, i.e., continuous and discontinuous, was accomplished to collect individually nanoHPLC-separated species within distinct spots and consequently recover by MALDI MS screening all major and minor GSL species in the mixtures. The nanoHPLC/MALDI MS coupling protocol was developed and applied to a mixture of neutral GSLs purified from human erythrocytes and a monosialoganglioside mixture expressed by the murine MDAY-D2 cell line. Additionally, on-line nanoHPLC/MALDI doping with lithium cations of individually separated neutral GSLs was introduced to enhance data interpretation of the GSL MS pattern, while preserving the same level of information and ultimately to enhance structural assignment of components of interest. The method is demonstrated to be highly sensitive, reaching the low femtomole level of detection of individual GSL species and is highlighted as a versatile analytical tool for glycolipidomic studies. Figure Automatic LC/MALDI MS profiling of glycosphingolipids Mostafa Zarei and Stephan Kirsch contributed equally to this work.     

A quantitative model of ultraviolet matrix-assisted laser desorption/ionization

A quantitative model of primary ionization in ultraviolet matrix-assisted laser desorption/ionization (UV-MALDI) is presented. It includes not only photochemical processes such as exciton pooling, but also the effects of the desorption event. The interplay of these two is found to be a crucial aspect of the MALDI process. The desorbing plume is modeled as an adiabatic expansion with entrained clusters. The parameters in the model are defined as much as possible via experiment or by analogy with known effects. The model was applied to the matrix 2,5-dihydroxybenzoic acid and found to reproduce the fluence dependence of the fluorescence yield and key features of the picosecond two-pulse ion generation efficiency curves. In addition, the model correctly predicts a fluence rather than irradiance threshold, the magnitude of the threshold, the magnitude of the ion yield, laser wavelength effects, plume temperatures, plume expansion velocities and the spot size effect.     

A novel approach of combining thin-layer chromatography with surface-assisted laser desorption/ionization (SALDI) time-of-flight mass spectrometry

A novel means of combining thin-layer chromatography (TLC) with laser desorption/ionization mass spectrometry using a liquid matrix is proposed. Surface-assisted laser desorption/ionization (SALDI) mass spectrometry, which uses a mixture of a micrometer-sized carbon powder (graphite or activated carbon, the SALDI solid) and 15% sucrose/glycerol, dissolved in an equal volume of methanol (SALDI liquid) as a SALDI matrix, is used for laser desorption mass analysis. The ablation of carbon powder from a pencil drawing was used as an alternative to the SALDI solid. The liquid matrix resembled that used in a conventional SALDI matrix system. A line was drawn before separation with a pencil on the track of the sample developed on the TLC plate. After TLC separation, approximately 0.1 microl of SALDI liquid was directly applied to the chromatographic spots on the TLC plate. Porphyrins were used to demonstrate this combination owing to the visible colors of this type of compound. The analyte signal can be easily detected by irradiating the laser along the pencil line on the TLC plate. An additive, p-toluenesulfonic acid, is added to the SALDI liquid to enhance the signal's intensity. This additive dramatically improves the signal-to-noise ratio. A detection limit of approximately 500 pg is demonstrated for porphines, which is 50 times better than that corresponding to conventional TLC SALDI.     

Two-laser infrared and ultraviolet matrix-assisted laser desorption/ionization

Matrix-assisted laser desorption/ionization (MALDI) was performed using two pulsed lasers with wavelengths in the IR and UV regions. A 10.6 micro m pulsed CO(2) laser was used to irradiate a MALDI target, followed after an adjustable delay by a 337 nm pulsed nitrogen laser. The sample consisted of a 2,5-dihydroxybenzoic acid matrix and bovine insulin guest molecule. The pulse energy for both of the lasers was adjusted so that the ion of interest, either the matrix or guest ion, was not produced by either of the lasers alone. The delay time for maximum ion yield occurs at 1 micro s for matrix and guest ions and the signal decayed to zero in approximately 400 micro s. A mechanism is presented for enhanced UV MALDI ion yield following the IR laser pulse based on transient heating.     

Multiphoton ionization spectroscopy in surface analysis and laser desorption mass spectrometry

The application of resonance-enhanced multiphoton ionization (REMPI) spectroscopy for the ultrasensitive detection of molecules originating from laser desorption experiments performed on a variety of substrates is reviewed. Laser-induced desorption from surfaces is capable of producing intact gas-phase molecules, even from polar, non-volatile, high-molecular-weight and thermally labile substances. REMPI is a highly efficient and optically selective ionization method, which, coupled with laser desorption allows the direct chemical analysis of complex mixtures, without the need for previous sample purification and separation steps. The use of REMPI spectroscopy is discussed in two contexts: (1) for the direct chemical analysis of complex mixtures, e.g., environmental samples, by laser desorption/laser postionization mass spectrometry and (2) for measurements of internal state distribution of molecules laser-desorbed from sub-monolayers surface films to gain insight into the laser desorption mechanism.Presented at the 13th International Symposium on Microchemical Techniques (ISM), held in Montreux, Switzerland, May 16–20,1994     

Decomposition of protein nitrosothiolsin matrix-assisted laser desorption/ionization and electrospray ionization mass spectrometry

The S-nitrosylation of proteins is involved in the trafficking of nitric oxide (NO) in intra- and extracellular milieus. To establish a mass spectrometric method for identifying this post-translational modification of proteins, a synthetic peptide and transthyretin were S-nitrosylated in vitro and analyzed by electrospray ionization (ESI) and matrix-assisted laser desorption/ionization (MALDI) mass spectrometry. The intact molecular ion species of nitrosylated compounds was identified in the ESI mass spectrum without elimination of the NO group. However, the labile nature of the S-NO bond was evident when the in-source fragmentation efficiently generated [M + H - 30](+) ions. The decomposition was prominent for multiply charged transthyretin ions with high charge states under ordinary ESI conditions, indicating that the application of minimum nozzle potentials was essential for delineating the stoichiometry of nitrosylation in proteins. With MALDI, the S-NO bond cleavage occurred during the ionization process, and the subsequent reduction generated [M + H - 29](+) ions.     

Characterization of some synthetic Ru and Ir complexes by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry

Matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOFMS) was applied to the analysis of Ru(OCOCF(3))(2)(CO)(PPh(3))(2), Ru(OCOC(3)F(7))(2)(CO)(PPh(3))(2), Ir(tBuppy)(3) and Ir(ppy)(2)(acac) complexes. A troublesome problem in the MALDI-TOFMS characterization of these metal complexes is the possible replacement of complex ligands by matrix. In this contribution, 10 matrices, ranging from acidic to basic, were investigated: alpha-cyano-4-hydroxycinnamic acid (CHCA), 2,5-dihydroxybenzoic acid (DHB), sinapinic acid (SA), dithranol, 2,4,6-trihydroxyactophenone (THAP), 6-azo-2-thiothymine (ATT), norharman, 2-[(2E)-3-(4-tert-butylphenyl)-2-methylprop-2-enylidene]malononitrile (DCTB), 4-nitroaniline (NA) and 2-amino-5-nitrophyridine (ANP). With most of the matrices, including the neutral and basic ones, matrix substitution of ligand could clearly be detected. Based on the experimental results, possible mechanisms of matrix substitution were discussed. It was demonstrated that the ligand exchange process might also occur through the gas-phase reactions initiated by laser shots. Among the matrices tested, DCTB was found to be the best one for the complexes that are prone to ligand exchange by matrix.     

Detection of water-soluble vitamins by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry using porphyrin matrices

The detection of water-soluble vitamins B(1), B(2), B(6), B(12) and C by matrix-assisted laser desorption/ionization (MALDI) time-of-flight mass spectrometry (TOFMS) was attempted by studying 17 porphyrin matrices. Comparative studies of porphyrin matrices, useful mass spectral window, matrix/analyte molar ratio (M/A), ultraviolet-visible absorption characteristics and quantitative results were made. Most porphyrin matrices provide a useful mass spectral window in the low-mass range. The optimal M/A increases with increasing molecular mass of the vitamin. Vitamin B(12) possesses the highest molecular mass and requires a higher M/A. The presence of hydroxyl or carboxyl groups in the porphyrin is an indicator of a useful MALDI matrix. Vitamins B(2) and B(6) were readily ionized upon irradiation with a 337 nm laser without the use of any porphyrin matrix. Improved linearity and sensitivity of the calibration curves were obtained with samples prepared with a constant M/A. The limits of detection and quantitation are at the picomole level. The results indicate that MALDI-TOFMS with porphyrin matrices is a rapid and viable technique for the detection of low molecular mass water-soluble vitamins.     

Identification of formylglycine in sulfatases by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry

C(alpha)-Formylglycine, the catalytic amino acid residue in the active site of sulfatases, is generated by post-translational modification of a cysteine or serine residue. We describe a highly sensitive procedure for the detection of C(alpha)-formylglycine-containing peptides in tryptic digests of sulfatase proteins. The protocol is based on the formation of hydrazone derivatives of C(alpha)-formylglycine-containing peptides when using dinitrophenylhydrazine as a matrix for matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOFMS). The hydrazone derivatives desorb and ionize with high efficiency and can be detected in the sub-femtomole range. The presence of C(alpha)-formylglycine is indicated by a mass increment of 180.13 u, corresponding to the hydrazone moiety, and also by a unique C-terminal fragment ion, characteristic of sulfatases, that becomes prominent in MALDI post-source decay mass spectra of the hydrazone derivatives.     

Fragmentation characteristics of peptide-metal ion adducts under matrix-assisted laser desorption/ionization post-source decay time-of-flight mass spectrometric conditions

Fragmentation reactions of sodium-cationized enkephalin peptides generated by matrix-assisted laser desorption/ionization were studied using post-source decay (PSD) with a reflectron time-of-flight mass spectrometer. Several matrices and analyte-matrix sample preparation methods were evaluated for high-intensity ion currents that could last for the entire PSD analysis. A triple dried-droplet sample preparation procedure with 2,5-dihydroxybenzoic acid as the matrix was found to yield abundant longer-lasting ion signals of the peptide-Na(+) ion adducts. The principal decay product of these adduct ions is the [b(n-1) + Na + OH](+) ion, which provides an unambiguous identification of the C-terminal residue of a peptide. In some peptides, the loss of a second residue from the C-terminus is also observed. No other sequence-specific ions were observed.     

Derivatization of small biomolecules for optimized matrix-assisted laser desorption/ionization mass spectrometry

Matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOFMS) is a powerful tool for the measurement of low molecular mass compounds of biological interest. The limitations for this method are the volatility of many analytes, possible interference with matrix signals or bad ionization or desorption behavior of the compounds. We investigated the application of well-known and straightforward one-pot derivatization procedures to circumvent these problems. The derivatizations tested allow the measurement and the labeling of alcohols, aldehydes and ketones, carboxylic acids, alpha-ketocarboxylic acids and amines.