Matrix-assisted Laser Desorption And Ionization Time-of-flight Research Articles

A mass spectrometry-based non-radioactive differential radial capillary action of ligand assay (DRaCALA) to assess ligand binding to proteins.

Binding of ligands to macromolecules changes their physicochemical and enzymatic characteristics. Cyclic di‐GMP is a second messenger involved in motility/sessility and acute/chronic infection life style transition. Although the GGDEF domain, predominantly a diguanylate cyclase, represents one of the most abundant bacterial domain superfamilies, the number of cyclic di‐GMP receptors falls short. To facilitate screening for cyclic di‐nucleotide binding proteins, we describe a non‐radioactive, matrix‐assisted laser desorption and ionization time‐of‐flight (MALDI‐TOF)‐based modification of the widely applied differential radial capillary action of ligand assay (DRaCALA). The results of this assay suggest that the diguanylate cyclase/phosphodiesterase variant YciRFec101, but not selected catalytic mutants, bind cyclic di‐GMP.Highlights Cyclic di‐nucleotides are ubiquitous second messengers in bacteria. However, few receptors have been identified.Previous screening of cell lysates by differential radial capillary action of ligand assay (DRaCALA) using radioactive ligand identified cyclic di‐nucleotide binding proteins.A MALDI‐TOF‐based DRaCALA was developed to detect cyclic di‐nucleotide binding as a non‐radioactive alternative.Known cyclic di‐GMP binding proteins were verified and potential cyclic di‐GMP binding proteins were identified.

Chemically heterogeneous carbon dots enhanced cholesterol detection by MALDI TOF mass spectrometry

A binary system composed of carbon dots (CDs) and N-doped CDs (N-CDs) embedded in an organic matrix was used for the analysis of cholesterol by MALDI (matrix-assisted laser desorption and ionization time-of-flight) mass spectrometry, as a model for detection of small, biologically relevant molecules. The results showed that both CDs are sensitive to the cholesterol and can be used either alone or in a binary system with 2,5-dihydroxybenzoic acid (DHB) to enhance the detection process. It was found that both COOH and NH2 groups on CDs surface contributed to the enhancement in the cholesterol detection by MALDI mass spectrometry in the presence of inorganic cations. Nevertheless, in the presence of NaCl, N-CDs led to a better reproducibility of results. It was due to the coexistence of positive and negative charge on N-CDs surface that led to a homogeneous analyte/substrate distribution, which is an important detection parameter. The enhancing effect of carbon dots was linked to a negative Gibbs energy of the complex formation between CDs, Na+, cholesterol and DHB, and it was supported by theoretical calculations. Moreover, upon the addition of CDs/N-CDs, such features as a low ionization potential, vertical excitation, dipole moment and oscillator strength positively affected the cholesterol detection by MALDI in the presence of Na+.

Proteomic Analysis of Tear Film Obtained from Diabetic Dogs.

Simple SummaryCanine diabetes is a serious disease, which can lead to severe complications, eventually even death. Currently, all the diagnostic procedures are the invasive ones, with blood collection remaining as a golden standard for both initial diagnosis, and later follow-up. Tears can be obtained in a non-invasive manner, which makes them a perfect candidate for a screening tool in canine diabetes. In this study we aimed to analyze the protein composition of the tears collected from the healthy animals and compared it to the diabetic group. There are significant differences between these two groups, and we believe that the identified proteins hold promise as a potential diagnostic tool, which can be later on used both in clinical practice, and for better understanding of the disease.Canine diabetes mellitus is a significant health burden, followed with numerous systemic complications, including diabetic cataracts and retinopathy, leading to blindness. Diabetes should be considered as a disease damaging all the body organs, including gastrointestinal tract, through a complex combination of vascular and metabolic pathologies, leading to impaired gut function. Tear film can be obtained in a non-invasive way, which makes it a feasible biomarker source. In this study we compared proteomic changes ongoing in tear film of diabetic dogs. The study group consisted of 15 diabetic dogs, and 13 dogs served as a control group. After obtaining tear film with Schirmer strips, we performed 2-dimensional electrophoresis, followed by Delta2D software analysis, which allowed to select statistically significant differentially expressed proteins. After their identification with MALDI-TOF (matrix assisted laser desorption and ionisation time of flight) spectrometry we found one up-regulated protein in tear film of diabetic dogs—SRC kinase signaling inhibitor 1 (SRCIN1). Eight proteins were down-regulated: phosphatidylinositol-4 kinase type 2 alpha (PI4KIIα), Pro-melanin concentrating hormone (Pro-MCH), Flotillin-1, Protein mono-ADP ribosyltransferase, GRIP and coiled coil domain containing protein 2, tetratricopeptide repeat protein 36, serpin, and Prelamin A/C. Identified proteins were analyzed by Panther Gene Ontology software, and their possible connections with diabetic etiopathology were discussed. We believe that this is the first study to target tear film proteome in canine diabetes. We believe that combined with traditional examination, the tear film proteomic analysis can be a new source of biomarkers both for clinical practice, and experimental research.

Geopolymer/CeO2 as Solid Electrolyte for IT-SOFC

As a material for application in the life sciences, a new composite material, geopolymer/CeO2 (GP_CeO2), was synthesized as a potential low-cost solid electrolyte for application in solid oxide fuel cells operating in intermediate temperature (IT-SOFC). The new materials were obtained from alkali-activated metakaolin (calcined clay) in the presence of CeO2 powders (x = 10%). Besides the commercial CeO2 powder, as a source of ceria, two differently synthesized CeO2 powders also were used: CeO2 synthesized by modified glycine nitrate procedure (MGNP) and self-propagating reaction at room temperature (SPRT). The structural, morphological, and electrical properties of pure and GP_CeO2-type samples were investigated by X-ray powder diffraction (XRPD), Fourier transform infrared (FTIR), BET, differential thermal and thermogravimetric analysis (DTA/TGA), scanning electron microscopy (FE-SEM), energy dispersive spectrometer (EDS), and method complex impedance (EIS). XRPD and matrix-assisted laser desorption and ionization time-of-flight (MALDI-TOF) analysis confirmed the formation of solid phase CeO2. The BET, DTA/TGA, FE-SEM, and EDS results indicated that particles of CeO2 were stabile interconnected and form a continuous conductive path, which was confirmed by the EIS method. The highest conductivity of 1.86 × 10−2 Ω−1 cm−1 was obtained for the sample GP_CeO2_MGNP at 700 °C. The corresponding value of activation energy for conductivity was 0.26 eV in the temperature range 500–700 °C.

Enhanced lysophosphatidylcholine and sphingomyelin contents are characteristic of spermatozoa from obese men—A MALDI mass spectrometric study

The lipids of human spermatozoa are characterized by unique fatty acyl compositions, i.e. considerable amounts of highly unsaturated, in particular docosahexaenoyl (22:6) fatty acyl residues. This makes spermatozoa very sensitive to oxidation. It has already been shown that the most abundant lipid of sperm, phosphatidylcholine (PC), is converted into lysophosphatidylcholine (LPC) under conditions of oxidative stress, and, thus, the PC/LPC ratio may be used as a measure of sperm quality. However, direct correlations between the PC/LPC ratios and certain pathologies are so far missing. We will show here for the first time (by using matrix-assisted laser desorption and ionization time-of-flight (MALDI-TOF) mass spectrometry (MS)) that sperm from obese donors (BMI>35kg/m2) are characterized by (a) a significantly increased LPC as well as (b) an increased sphingomyelin content. This does exclusively hold true for the lipid extracts of the spermatozoa but is not valid for the related seminal fluids.

IN-SITU PROBING OF RADIATION-INDUCED PROCESSING OF ORGANICS IN ASTROPHYSICAL ICE ANALOGS—NOVEL LASER DESORPTION LASER IONIZATION TIME-OF-FLIGHT MASS SPECTROSCOPIC STUDIES

Understanding the evolution of organic molecules in ice grains in the interstellar medium (ISM) under cosmic rays, stellar radiation, and local electrons and ions is critical to our understanding of the connection between ISM and solar systems. Our study is aimed at reaching this goal of looking directly into radiation-induced processing in these ice grains. We developed a two-color laser-desorption laser-ionization time-of-flight mass spectroscopic method (2C-MALDI-TOF), similar to matrix-assisted laser desorption and ionization time-of-flight (MALDI-TOF) mass spectroscopy. Results presented here with polycyclic aromatic hydrocarbon (PAH) probe molecules embedded in water-ice at 5 K show for the first time that hydrogenation and oxygenation are the primary chemical reactions that occur in astrophysical ice analogs when subjected to Lyα radiation. We found that hydrogenation can occur over several unsaturated bonds and the product distribution corresponds to their stabilities. Multiple hydrogenation efficiency is found to be higher at higher temperatures (100 K) compared to 5 K—close to the interstellar ice temperatures. Hydroxylation is shown to have similar efficiencies at 5 K or 100 K, indicating that addition of O atoms or OH radicals to pre-ionized PAHs is a barrierless process. These studies—the first glimpses into interstellar ice chemistry through analog studies—show that once accreted onto ice grains PAHs lose their PAH spectroscopic signatures through radiation chemistry, which could be one of the reason for the lack of PAH detection in interstellar ice grains, particularly the outer regions of cold, dense clouds or the upper molecular layers of protoplanetary disks.

2,4-Dinitrophenylhydrazine as a New Reactive Matrix to Analyze Oxidized Phospholipids by MALDI-TOF Mass Spectrometry

Matrix-assisted laser desorption and ionization time-of-flight (MALDI-TOF) mass spectrometry (MS) is increasingly used to analyze native and oxidized lipids. However, as the majority of MALDI devices are operating under high vacuum, the detection of volatile compounds such as “small” aldehydes is difficult and requires previous derivatization. This study uses 2,4-dinitrophenylhydrazine (DNPH) to efficiently convert aldehydes into hydrazones that can be easily identified by the characteristic mass shift and their selective enrichment by precipitation. Additionally, the UV absorption of the hydrazones at 337 nm is sufficiently high allowing direct analysis without typical MALDI matrices. Finally, it will be shown that the presence of DNPH does also enable the detection of non-oxidized phospholipids.

Parameters affecting the accuracy of the MALDI-TOF MS determination of the phosphatidylcholine/lysophosphatidylcholine (PC/LPC) ratio as potential marker of spermatozoa quality

Matrix-assisted laser desorption and ionization time-of-flight (MALDI-TOF) mass spectrometry (MS) is increasingly used to characterize (phospho)lipids. However, quantitative MALDI data are often questioned because ion suppression may occur if mixtures are analyzed. Therefore, relative (but no absolute) data are normally derived from the MALDI mass spectra of lipid mixtures. We are particularly interested in the phosphatidylcholine/lysophosphatidylcholine (PC/LPC) ratio because it seems to represent a suitable measure of the inflammatory activity. In this study, different parameters affecting the achievable accuracy of the MALDI-TOF MS determination of the PC/LPC ratio are compared. It will be shown that particularly the applied laser fluence as well as the used solvents influence the accuracies. Using artificial lipid mixtures it will be demonstrated that the PC/LPC ratio can be determined with an accuracy of about ±10% making the MALDI assay comparable to established methods. Finally, it will be shown that the optimized conditions are also useful to determine the PC/LPC ratios in human seminal plasma.

Global proteomic analysis of plasma from mice infected with Plasmodium berghei ANKA using two dimensional gel electrophoresis and matrix assisted laser desorption ionization-time of flight mass spectrometry

BackgroundA global proteomic strategy was used to identify proteins, which are differentially expressed in the murine model of severe malaria in the hope of facilitating future development of novel diagnostic, disease monitoring and treatment strategies.MethodsMice (4-week-old CD1 male mice) were infected with Plasmodium berghei ANKA strain, and infection allowed to establish until a parasitaemia of 30% was attained. Total plasma and albumin depleted plasma samples from infected and control (non-infected) mice were separated by two-dimensional gel electrophoresis (2-DE). After staining, the gels were imaged and differential protein expression patterns were interrogated using image analysis software. Spots of interest were then digested using trypsin and the proteins identified using matrix-assisted laser desorption and ionization-time of flight (MALDI-TOF) mass spectrometry (MS) and peptide mass fingerprinting software.ResultsMaster gels of control and infected mice, and the corresponding albumin depleted fractions exhibited distinctly different 2D patterns comparing control and infected plasma, respectively. A wide range of proteins demonstrated altered expression including; acute inflammatory proteins, transporters, binding proteins, protease inhibitors, enzymes, cytokines, hormones, and channel/receptor-derived proteins.ConclusionsMalaria-infection in mice results in a wide perturbation of the host serum proteome involving a range of proteins and functions. Of particular interest is the increased secretion of anti-inflammatory and anti apoptotic proteins.

A simple TLC-MALDI method to monitor oxidation products of phosphatidylcholines and -ethanolamines

Summary The analysis of lipid-phospholipid oxidation products is of primary importance. Although there are established HPLC and LC-MS techniques, it is shown here for the first time that the combination of matrix-assisted laser desorption and ionization time-of-flight (MALDI-TOF) mass spectrometry (MS) and TLC represents a sensitive, fast, and convenient alternative. A mixture of 1-palmitoyl-2-linoleoyl-sn-phosphatidylcholine (PLPC) and -ethanolamine (PLPE) was oxidized under the influence of atmospheric oxygen and characterized by direct positive ion MALDI-TOF MS as well as combined TLC-MALDI. It is shown that much more detailed information — particularly related to the oxidation products of PLPE that have so far been scarcely investigated — can be obtained by TLC-MALDI. However, it is also shown that further methodological improvements are necessary to make this method generally applicable to complex lipid mixtures.

Comparison of MALDI-TOF mass spectra of [PdCl(dien)]Cl and [Ru(en)2Cl2]Cl acquired with different matrices

In this work, the matrix-assisted laser desorption and ionization time-of-flight (MALDI-TOF) mass spectra of two cationic complexes, i.e., [PdCl(dien)]Cl and [Ru(en)2Cl2]Cl, acquired under different conditions were analyzed. The spectra were recorded with three matrices with or without trifluoroacetic acid (TFA), i.e., two traditional matrices, i.e., 2,5-dihydroxybenzoic acid and ?-cyano-hydroxycinnamic acid, and one flavonoid, quercetin. The spectra acquired with quercetin appeared to be the simplest, whereas in the spectra obtained with other matrices, peaks arising either from the addition of matrix molecules or from the fragmentation products were detectable. Addition of TFA did not complicate the spectra of the Pd(II) and Ru(III) complexes when the traditional matrices were used. On the other hand, the spectra of Pd complex were simpler, whereas the addition of TFA in the case of the Ru complex resulted in a higher number of peaks, some of which could not be identified. Taken together, the results of this study once more emphasize the differences arising in the MALDI-TOF mass spectra of transition metal complexes in dependence on the applied matrix.

Novel biobased epoxy compounds: epoxidized sucrose esters of fatty acids

Novel biobased high functionality epoxy resins were synthesized by the epoxidation of sucrose ester resins of vegetable oil fatty acids (SEFA). A series of sucrose esters of fatty acids were epoxidized by peracetic acid generated in situ from hydrogen peroxide and acetic acid in the presence of an ion exchange resin catalyst to produce the epoxidized sucrose esters of fatty acids (ESEFA). The conversion of double bonds to epoxides was greater than 99%. The products were characterized by matrix-assisted laser desorption and ionization time-of-flight (MALDI-TOF) mass spectrometry, Fourier-transform infrared (FTIR) spectroscopy, and nuclear magnetic resonance (NMR) spectroscopy. The physical properties were characterized using bulk viscosity, intrinsic viscosity, and density measurements. The thermal properties were characterized by differential scanning calorimetry (DSC) and thermogravimetric analysis (TGA). Since sucrose esters consist of a sucrose core fully substituted with eight vegetable oil fatty acids, both SEFAs and ESEFAs possess well-defined compact macromolecular structures. As novel biobased epoxy compounds, ESEFAs possess high epoxy functionality (8–15 per molecule), high density, and a compact molecular structure. Thus, ESEFAs are very promising in a number of uses from the formation of biobased thermosets by crosslinking of the epoxy groups, to further derivatization by reaction of the epoxy groups.

Cathepsin X is secreted by human osteoblasts, digests CXCL-12 and impairs adhesion of hematopoietic stem and progenitor cells to osteoblasts

Hematopoietic stem cells are retained within discrete bone marrow niches through the effects of cell adhesion molecules and chemokine gradients. However, a small proportion of hematopoietic stem cells can also be found trafficking in the peripheral blood. During induced stem cell mobilization a proteolytic microenvironment is generated, but whether proteases are also involved in physiological trafficking of hematopoietic stem cells is not known. In the present study we examined the expression, secretion and function of the cysteine protease cathepsin X by cells of the human bone marrow. Human osteoblasts, bone marrow stromal cells and hematopoietic stem and progenitor cells were analyzed for the secretion of cathepsin X by western blotting, active site labeling, immunofluorescence staining and activity assays. A possible involvement of cathepsin X in cell adhesion and CXCL-12-mediated cell migration was studied in functional assays. Matrix-assisted laser desorption and ionization time-of-flight (MALDI-TOF) analysis revealed the digestion mechanism of CXCL-12 by cathepsin X. Osteoblasts and stromal cells secrete cathepsin X, whereas hematopoietic stem and progenitor cells do not. Using a cathepsin X-selective substrate, we detected the catalytic activity of cathepsin X in cell culture supernatants of osteoblasts. Activated cathepsin X is able to reduce cellular adhesive interactions between CD34(+) hematopoietic stem and progenitor cells and adherent osteoblasts. The chemokine CXCL-12, a highly potent chemoattractant for hematopoietic stem cells secreted by osteoblasts, is readily digested by cathepsin X. The exo-peptidase cathepsin X has been identified as a new member of the group of CXCL-12-degrading enzymes secreted by non-hematopoietic bone marrow cells. Functional data indicate that cathepsin X can influence hematopoietic stem and progenitor cell trafficking in the bone marrow.

Significant sensitivity improvements by matrix optimization: a MALDI-TOF mass spectrometric study of lipids from hen egg yolk

Due to its sensitivity, the tolerance of impurities and the simplicity of performance, matrix-assisted laser desorption and ionization time-of-flight (MALDI-TOF) mass spectrometry (MS) is increasingly used to analyze lipids from biological sources. Although its detailed role is not understood so far, the applied matrix has a pronounced effect on the achievable spectrum quality and particularly how sensitive the individual lipid classes are detectable. Different matrix compounds were recently established in the lipid field including 2,5-dihydroxybenzoic acid (DHB), 9-aminoacridine (9-AA), para-nitroaniline (PNA), 2-mercaptobenzothiazole (MBT), and 2-(2-aminoethylamino)-5-nitropyridine (AAN). It is the aim of this paper to compare the properties of these matrices with the newly synthesized matrix, α-cyano-2,4-difluorocinnamic acid (Di-FCCA). An organic extract from hen egg yolk was used as a simple and easily available test system. It will be shown that Di-FCCA is the matrix of choice to detect lipids in the positive-ion mode due to an achievable sensitivity gain of more than one order of magnitude compared to alternative matrices. In contrast, Di-FCCA is not suitable for negative-ion detection of phospholipids. Here, 9-AA is unequivocally the matrix of choice.

Differently complex oligosaccharides can be easily identified by matrix-assisted laser desorption and ionization time-of-flight mass spectrometry directly from a standard thin-layer chromatography plate

Thin-layer chromatography (TLC) is a simple, fast and inexpensive separation method. Unambiguous identification of the TLC spots is, however, often a problem. Here we show for the first time that oligosaccharides (derived from dextran, alginate, hyaluronan and chondroitin sulfate) can be characterized by matrix-assisted laser desorption and ionization time-of-flight (MALDI-TOF) mass spectrometry (MS) directly on a TLC plate. The applied oligosaccharides were either commercially available or obtained from the polysaccharides by HCl-induced hydrolysis. Normal phase TLC was followed by MALDI-TOF MS subsequent to matrix deposition. It will be shown that high quality mass spectra can be obtained that enable unequivocal assignments. It will also be shown that the high content of formic acid in the solvent system does not confer major problems but is responsible for the partial formylation of the analyte and minor N-acetyl loss from hyaluronan and chondroitin sulfate.

The intact muscle lipid composition of bulls: an investigation by MALDI-TOF MS and 31P NMR

The analysis of beef lipids is normally based on chromatographic techniques and/or gas chromatography in combination with mass spectrometry (GC/MS). Modern techniques of soft-ionization MS were so far scarcely used to investigate the intact lipids in muscle tissues of beef. The objective of the study was to investigate whether matrix-assisted laser desorption and ionization time-of-flight (MALDI-TOF) mass spectrometry and (31)P nuclear magnetic resonance (NMR) spectroscopy are useful tools to study the intact lipid composition of beef. For the MALDI-TOF MS and (31)P NMR investigations muscle samples were selected from a feeding experiment with German Simmental bulls fed different diets. Beside the triacylglycerols (TAGs), phosphatidylethanolamine (PE), phosphatidylcholine (PC) and phosphatidylinositol (PI) species the MALDI-TOF mass spectra of total muscle lipids gave also intense signals of cardiolipin (CL) species. The application of different matrix compounds, 2,5-dihydroxybenzoic acid (DHB) and 9-aminoacridine (9-AA), leads to completely different mass spectra: 9-AA is particularly useful for the detection of (polar) phospholipids, whereas apolar lipids, such as cholesterol and triacylglycerols, are exclusively detected if DHB is used. Finally, the quality of the negative ion mass spectra is much higher if 9-AA is used.

Matrix-assisted laser desorption and ionisation time-of-flight mass spectrometry of Pt(II) and Pd(II) complexes

In this work, we have analysed matrix-assisted laser desorption and ionisation time-of-flight (MALDI-TOF) mass spectra of [PtCl 2( en)], [PtCl 2( dach)] and [PdCl( dien)]Cl acquired either with 2,5-dihydroxybenzoic acid (DHB) or α-cyano-hydroxycinnamic acid (CHCA) as matrices. For certain experiments, small amounts of trifluoro acetic acid (TFA) or higher concentration of inorganic salts (NaCl or KCl) was added to the matrix solution. The majority of peaks arising from the Pt(II) and Pd(II) complexes could be identified, but certain ions detectable in the spectra were generated upon ligand loss. Additionally, the analysis of Pt(II) complexes was also possible in the presence of a higher salt content, which is a commonly used analysis condition for the samples of biological origin. While DHB appears to be the best suited for MALDI-TOF mass spectrometric analysis of Pt(II) complexes, CHCA seems to be a better matrix for Pd(II) complex used in this study. On the other hand, small amounts of TFA improve the spectra quality of Pt(II) complexes, but lead most probably to the degradation of Pd(II) complex. Taken together, we have demonstrated that the analysis of metallo-drugs using MALDI-TOF MS, though accompanied with certain identification problems, is easy and reliable. On the other hand, having in mind that some complexes (i.e. a combination of a particular transition metal/ligand) cannot be analysed under conditions usually applied for others, we deem it necessary to find out the best conditions for MALDI-TOF MS analysis of each metal complex.

Application of MALDI‐TOF mass spectrometry in lipidomics

AbstractLipids are of high interest on different levels. First, lipids as triacylglycerols (TAG) (particularly different vegetable oils) are of immense commercial interest. Second, lipids are of high relevance for different in vivo processes. For instance, the diagnostics of changes of the lipoproteins of human blood is very important in vascular diseases, whereas phospholipids (PL) are crucial for the properties of the cellular membrane and some selected PL were also recognized to represent important second messengers. Although there are many different methods (including different chromatographic and spectroscopic approaches) available, mass spectrometry (MS) is the most versatile tool of lipid analysis, with or without prior separation steps. Although several different ionization techniques are capable of producing ions from the lipids of interest, matrix‐assisted laser desorption and ionization time‐of‐flight (MALDI‐TOF) MS is the simplest and most convenient method. This review gives a short methodological survey of the capabilities and drawbacks of MALDI‐TOF MS and discusses its role in lipidomic studies. Although a summary about PL analysis will also be given, the focus of this review is the analysis of apolar lipids as TAG, cholesterol and cholesteryl esters. Aspects of sample preparations and the avoidance of pitfalls are also highlighted. Finally, it is the aim of this paper to show that MALDI MS provides data comparable to other modern MS methods, but in a faster and more convenient way.

Alignment of protein mass spectrometry data by integrated Markov chain shifting method

Mass spectrometers such as SELDI-TOF (surface enhanced laser desorption/ionization time-of-flight) and MALDI-TOF (matrix assisted laser desorption and ionization time-of-flight) measure the relative abundance of different protein ions or protein fragments (peptides) indexed by the mass-to-charge ratio (m/z). A special characteristic of the MS spectra is its variabilities in both m/z values and intensity magnitudes. We propose modelling the logintensities by a semiparametric model and the m/z by the integrated Markov chain shifting (IMS) model, for which the second-order differences of the random effects are assumed to follow a second-order Markov chain. Alignment of spectra is done through averaging over the random shifts conditional on the observed intensity information. The unknown parameters are estimated by an iterative nonparametric maximum profile likelihood method and a Gaussian kernel approximation. The bandwidths in kernel approximation are taken to be 0.04%–0.08% of the m/z values. Simulation results show that the proposed approach can achieve satisfactory alignment by reducing the intensity variations of the misalignment spectra by a factor of around 75%. Most alignment algorithms align spectra by clustering neighboring peaks and do not incorporate peak height information. Our semiparametric random shifting method builds a model taking into consideration of both the random shift effects of neighboring m/z values and similarity of the intensity magnitudes of common peaks within the ranges of about 50% of the intensity values.

MALDI-TOF “fingerprint” phospholipid mass spectra allow the differentiation between ruminantia and feloideae spermatozoa

The detailed comparative analysis of sperm lipids could essentially contribute to a better understanding of membrane function in the context of fertilization and, moreover, of sperm preservation. The application of sensitive analytical methods is particularly necessary for endangered species as the available amount of spermatozoa (and, accordingly, extractable lipids) is strongly limited. It will be shown that matrix-assisted laser desorption and ionization time-of-flight (MALDI-TOF) mass spectrometry (MS) is a fast, simple and sensitive method for the determination of the phospholipid composition of spermatozoa from several ruminantia (cattle, roe deer, Klipspringer) and feloideae species (domestic cat, Siberian tiger, fosa). Characteristic “fingerprints” are obtained from the positive ion spectra that allow the differentiation between both animal groups. In contrast to the lipid extracts of ruminantia spermatozoa which predominantly contain ether lipids including essential amounts of plasmalogens, the more complex phospholipid composition of feloideae spermatozoa is clearly dominated by diacyl phospholipids and contains only marginal amounts of plasmalogens. It will also be shown that the lipid compositions of ejaculated, electroejaculated and cauda epididymal spermatozoa of the same species are very similar and give comparable data. Therefore, the analysis of ejaculated spermatozoa is not an absolute must.