Figure 2
(a) MALDI-TOF peptide profile obtained from human serum after sample workup using RPC18-functionalized magnetic beads. (b) Observed isotopic distributions of five peptides chosen for alignment of all serum peptide profiles. The horizontal lines at the top of the peaks represent the calculated values of the corresponding polyaveragine peptides. (c) Profile processing results in a QC value for each individual mass spectrum. First, the intensity of each isotope peak is determined using Xtractor. Then, after normalization, the observed intensities are compared with those estimated from the polyaveragine peptide. For a more accurate determination of the deviation, the random noise variance is taken into account. Finally, the sum of thus obtained deviations for all considered peptides represents the QC value of the MALDI-TOF peptide profile in (a).
Source publication
In this study, we have implemented a new quality control (QC) parameter for peptide profiling based on isotopic distributions. This QC parameter is an objective measure and facilitates automatic sorting of large numbers of peptide spectra. Peptides in human serum samples were enriched using reversed-phase C(18)-functionalized magnetic beads using a...
Contexts in source publication
Context 1
... of the aims of this study was to demonstrate the use of peptide isotopic distri- butions of peptides for quality control parameter. For illustration, the isotopic patterns of the five ubiquitous peptides used for alignment are shown in Figure 2. While the selection of these five peptides was an arbitrary process, no software is available that yields a better result without the need of user-defined input parameters. ...
Context 2
... is sufficient for the estimation of isotopic distribution of unknown peptides since we are not searching for the best fit, but only need to discriminate between peptide and non- peptide (e.g., polymer, matrix cluster) distributions. The estimated values of the five peptides are depicted in Figure 2. These are obtained by first transferring a peptide m/z value into a neutral mass (i.e., [m/z value- proton-water]), then dividing this number by the exact mass of averagine (i.e., 111.0543 ...
Context 3
... to rounding of the indices, the m/z values of the observed peptide and the corresponding polyaveragine are not identical. It is clear from Figure 2 that the isotopic patterns of the five peptides used for alignment were in good agreement with the polyaver- agine distributions. A quantitative evaluation of these isotope distributions will be given in the next section. ...
Context 4
... intensities of the observed peaks in each peptide isotope distribution were determined using Xtractor (see the Materials and Methods section). As an example, for the peptide at m/z 1865.22 the intensity of the monoisotopic was determined as the sum of all mea- sured values at m/z 1865.22 0.49, the intensity of the second isotope was determined as the sum of all measured values at m/z 1866.22 0.49, and so forth (Figure 2). The window of 0.49 Da was chosen to ensure total quantification of each isotope. ...
Context 5
... and false-negative results. For a quantita- tive analysis, the normalized intensities estimated from the polyaveragine peptides, the average ratios of the isotopic distributions and their coefficients of variation were evaluated for the five peptides discussed earlier (see Figure 2) and the results are summarized in Table 1. In this evaluation, the 50 MALDI-TOF and the 50 MALDI-FTICR mass spectra with lowest QC values were selected from the results obtained from 192 pep- tide profiles in Figure 5. ...
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Due to their role in many diseases, enzymes of the ubiquitin system have recently become interesting drug targets. Despite efforts, primary screenings of compound libraries targeting E2 enzymes and E3 ligases have been strongly limited by the lack of robust and fast high-throughput assays. Here we report a label-free high-throughput screening assay...
Citations
... One of the advantages of using ultrahigh-resolution MS is that measurements at isotopic resolution provide more spectra information compared to broad-peak detection in linear mode MALDI-TOF MS. Previously, we showed that the goodness of the observed isotopic distributions can be used as a quality control parameter for the selection of high-quality spectra generated from the analysis of a large cohort of samples (Nicolardi et al., 2010). This concept was then implemented in a more powerful software-namely, MassyTools-developed for the high-throughput processing of MALDI mass spectra (Jansen et al., 2015). ...
Apolipoprotein-CIII (apo-CIII) is a glycoprotein involved in lipid metabolism and its levels are associated with cardiovascular disease risk. Apo-CIII sialylation is associated with improved plasma triglyceride levels and its glycosylation may have an effect on the clearance of triglyceride-rich lipoproteins by directing these particles to different metabolic pathways. Large-scale sample cohort studies are required to fully elucidate the role of apo-CIII glycosylation in lipid metabolism and associated cardiovascular disease. In this study, we revisited a high-throughput workflow for the analysis of intact apo-CIII by ultrahigh-resolution MALDI FT-ICR MS. The workflow includes a chemical oxidation step to reduce methionine oxidation heterogeneity and spectrum complexity. Sinapinic acid matrix was used to minimize the loss of sialic acids upon MALDI. MassyTools software was used to standardize and automate MS data processing and quality control. This method was applied on 771 plasma samples from individuals without diabetes allowing for an evaluation of the expression levels of apo-CIII glycoforms against a panel of lipid biomarkers demonstrating the validity of the method. Our study supports the hypothesis that triglyceride clearance may be regulated, or at least strongly influenced by apo-CIII sialylation. Interestingly, the association of apo-CIII glycoforms with triglyceride levels was found to be largely independent of body mass index. Due to its precision and throughput, the new workflow will allow studying the role of apo-CIII in the regulation of lipid metabolism in various disease settings.
... 39 Deviations from the theoretical isotopic distribution may be indicative of deviations in the elemental composition or the presence of overlapping ion species. 40 Fourier transform ion cyclotron resonance (FT-ICR) MS provides higher resolving power than TOF MS. 41 FT-ICR MS measurements of cesium iodide clusters [Cs(CsI) n ] + , up to about m/z 32,000, have been previously reported. 42,43 Ultrahigh-resolution MALDI FT-ICR MS has been used to analyze biomolecules at an isotopic resolution up to about m/z 24,000 44−47 and characterize complex biomolecules such as monoclonal antibodies. ...
Carbohydrates, such as oligo- and polysaccharides, are highly abundant biopolymers that are involved in numerous processes. The study of their structure and functions is commonly based on a material that is isolated from complex natural sources. However, a more precise analysis requires pure compounds with well-defined structures that can be obtained from chemical or enzymatic syntheses. Novel synthetic strategies have increased the accessibility of larger monodisperse polysaccharides, posing a challenge to the analytical methods used for their molecular characterization. Here, we present wide mass range ultrahigh-resolution matrix-assisted laser desorption/ionization (MALDI) Fourier transform ion cyclotron resonance (FT-ICR) mass spectrometry (MS) as a powerful platform for the analysis of synthetic oligo- and polysaccharides. Synthetic carbohydrates 16-, 64-, 100-, and 151-mers were mass analyzed and characterized by MALDI in-source decay FT-ICR MS. Detection of fragment ions generated from glycosidic bond cleavage (or cross-ring cleavage) provided information of the monosaccharide content and the linkage type, allowing for the corroboration of the carbohydrate compositions and structures.
... The molecular isotopic distribution measured by mass spectrometry contains elemental composition, facilitating molecular composition determination [28]. It has been demonstrated that the quality of peptide profiling can be improved by the comparison between the measured isotope patterns and the calculated ones [29]. In Fig (4B-4E), we compared the observed isotopic patterns of molecules to the theoretical ones calculated by the IsotopePattern software. ...
Background:
Frankincense is a resin secreted by the Boswellia tree. It is used in perfumery, aromatherapy, skincare, and traditional Chinese medicine. However, all Boswellia species are under threat owing to habitat loss and overexploitation. As a result, the market is getting flooded with counterfeit frankincense products.
Objective:
This study aims to establish a high-throughput method to screen and identify the authenticity of commercial frankincense products. We report, for the first time, a matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF MS)-based method for rapid and high-throughput screening of frankincense samples.
Methods:
MALDI-TOF MS, HPLC, thin-layer chromatography (TLC), and in vitro anti-inflammatory activity assay were used to examine the frankincense samples.
Results:
Well-resolved peaks of frankincense triterpenoids in the spectra were observed in the crude extract of commercial samples, including α-boswellic acids (αBAs), β-boswellic acids (βBAs), 11-keto-β-boswellic acids (KBAs), acetyl-11-keto-β-boswellic acids (AKBAs), and their esters. These compounds can be used as indicators for determining the authenticity of frankincense.
Conclusion:
Unlike LC-MS, which is a time-consuming and expensive method, and TLC, which requires a reference sample, our inexpensive, rapid high-throughput identification method based on MALDI-TOF MS is ideal for large-scale screening of frankincense samples sold in the market.
... The presence of organic deposits on the ZnO NCs' surfaces was studied by both spectroscopic (FT-IR) ( Figure 3) and spectrometric (LDI-MS) ( Figure 4) techniques. Fourier-transform infrared spectroscopy proved the presence of functional groups which contributed to the process of nano-ZnO formation, while mass spectrometry is a useful technique for the analysis of metal/metal oxide nanomaterials with organic deposits, mainly due to the isotopic pattern of d-electron metals [42]. Figure 3 represents the biologically synthesized ZnO NC spectra in the mid-infrared (MIR) range. ...
... The presence of organic deposits on the ZnO NCs' surfaces was studied by both spectroscopic (FT-IR) ( Figure 3) and spectrometric (LDI-MS) (Figure 4) techniques. Fourier-transform infrared spectroscopy proved the presence of functional groups which contributed to the process of nano-ZnO formation, while mass spectrometry is a useful technique for the analysis of metal/metal oxide nanomaterials with organic deposits, mainly due to the isotopic pattern of d-electron metals [42]. Figure 3 represents the biologically synthesized ZnO NC spectra in the mid-infrared (MIR) range. ...
This research presents, for the first time, the potential of the Lactobacillus paracasei LC20 isolated from sweet whey as a novel, effective and accessible source for post-cultured ZnO nanocomposites synthesis. The obtained nanocomposites were subjected to comprehensive characterization by a broad spectrum of instrumental techniques. Results of spectroscopic and microscopic analysis confirmed the hexagonal crystalline structure of ZnO in the nanometer size. The dispersion stability of the obtained nanocomposites was determined based on the zeta potential (ZP) measurements-the average ZP value was found to be −29.15 ± 1.05 mV in the 7-9 pH range. The ZnO nanocomposites (NCs) demonstrated thermal stability up to 130 • C based on the results of thermogravimetric TGA/DTG) analysis. The organic deposit on the nanoparticle surface was recorded by spectroscopic analysis in the infrared range (FT-IR). Results of the spectrometric study exhibited nanostructure-assisted laser desorption/ionization effects and also pointed out the presence of organic deposits and, what is more, allowed us to identify the specific amino acids and peptides present on the ZnO NCs surfaces. In this context, mass spectrometry (MS) data confirmed the nano-ZnO formation mechanism. Moreover, fluorescence data showed an increase in fluorescence signal in the presence of nanocomposites designed for potential use as, e.g., biosensors. Despite ZnO NCs' luminescent properties, they can also act as promising antiseptic agents against clinically relevant pathogens. Therefore, a pilot study on the antibacterial activity of biologically synthesized ZnO NCs was carried out against four strains (Escherichia coli, Staphylococcus aureus, Klebsiella pneumoniae and Pseudomonas aeruginosa) by using MIC (minimal inhibitory concentration). Additionally, the colony forming units (CFU) assay was performed and quantified for all bacterial cells as the percentage of viable cells in comparison to a control sample (untreated culture) The nanocomposites were effective among three pathogens with MIC values in the range of 86.25-172.5 µg/mL and showed potential as a new type of, e.g., medical path or ointment formulation.
... Therefore, OS extracted from the human milk sample were analyzed without the addition of the isotope-labeled standard (Figure 3(A)). The zoom into the m/z range from 726 to 746 (Figure 3(A-ii)) shows not only the expected isotopic distribution of tetraose (monoisotopic peak indicated by an asterisk) [85] but also an otherwise flat baseline. These findings approve the eligibility of [ 13 C6/ 15 Figure S7). ...
Several health benefits, associated with human milk oligosaccharides (HMOS), have been revealed in the last decades. Further progress, however, requires not only the establishment of a simple "routine" method for absolute quantification of complex HMOS mixtures but also the development of novel synthesis strategies to improve access to tailored HMOS. Here, we introduce a combination of salvage-like nucleotide sugar-producing enzyme cascades with Leloir-glycosyltransferases in a sequential pattern for the convenient tailoring of stable isotope-labeled HMOS. We demonstrate the assembly of [13C6]galactose into lacto-N- and lacto-N-neo-type HMOS structures up to octaoses. Further, we present the enzymatic production of UDP-[15N]GlcNAc and its application for the enzymatic synthesis of [13C6/15N]lacto-N-neo-tetraose for the first time. An exemplary application was selected—analysis of tetraose in complex biological mixtures—to show the potential of tailored stable isotope reference standards for the mass spectrometry-based quantification, using matrix-assisted laser desorption/ionization-time of flight mass spectrometry (MALDI-TOF-MS) as a fast and straightforward method for absolute quantification of HMOS. Together with the newly available well-defined tailored isotopic HMOS, this can make a crucial contribution to prospective research aiming for a more profound understanding of HMOS structure-function relations.
... One-dimensional and two-dimensional MALDI MS have been used for protein identification, sequencing, and searching for posttranslational modifications such as phosphorylation in casein components [13]. This strategy and knowledge of the isotopic pattern of d-electron metals allows the identification of organics binding to ions and clusters of metals such as metal proteins, metallopeptidases, and also metal and metal oxide nanoparticles [27]. Many researchers reported the identification of zinc ions bound to organics and chemically synthesized titanium oxide nanoclusters [28,29]. ...
Asymmetric flow field-flow fractionation coupled with use of ultraviolet–visible, multiangle light scattering (MALLS), and dynamic light scattering (DLS) detectors was used for separation and characterization of biologically synthesized silver composites in two liquid compositions. Moreover, to supplement the DLS/MALLS information, various complementary techniques such as transmission electron spectroscopy, Fourier transform infrared spectroscopy, and matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF MS) were used. The hydrodynamic diameter and the radius of gyration of silver composites were slightly larger than the sizes obtained by transmission electron microscopy (TEM). Moreover, the TEM results revealed the presence of silver clusters and even several morphologies, including multitwinned. Additionally, MALDI-TOF MS examination showed that the particles have an uncommon cluster structure. It can be described as being composed of two or more silver clusters. The organic surface of the nanoparticles can modify their dispersion. We demonstrated that the variation of the silver surface coating directly influenced the migration rate of biologically synthesized silver composites. Moreover, this study proves that the fractionation mechanism of silver biocolloids relies not only on the particle size but also on the type and mass of the surface coatings. Because silver nanoparticles typically have size-dependent cytotoxicity, this behavior is particularly relevant for biomedical applications.
Graphical abstractWorkflow for asymmetric flow field-flow fractionation of natural biologically synthesized silver nanocomposites
Electronic supplementary material
The online version of this article (10.1007/s00216-018-0967-0) contains supplementary material, which is available to authorized users.
... Currently, MALDI-TOF MS profiling of whole bacterial cells is being used more frequently for bacterial identification (Dec et al. 2014). Furthermore, based on shape of isotopic pattern of delectrons metals, it is possible to identify the organic compounds connected with these metals and clusters (Nicolardi et al. 2010). In this work, there were observed the signals of silver isotopes [ 107 Ag] + m/z 106.905 and [ 109 Ag] + m/z 108.910. ...
This research develops a safe, inexpensive, and more accessible source for synthesis of silver nanoparticles. The bioactive silver composites synthesized by Lactococcus lactis 56 KY484989 (LCLB56-AgCs) were characterized by various physico-chemical techniques and investigated for their antimicrobial activity and cytotoxicity. The average amount of nanoparticles was 0.363 ± 0.09 mg from 50 mL of culture medium. The synthesis efficiency varied from 71 to 85%. Synthesized silver nanoparticles with spherical in shape were found to be of 5–50 nm and average diameter 19 ± 2 nm. Based on the shape of isotopic pattern of d-electrons metals, the signals of silver isotopes [107Ag]+ at m/z 106.905 and [109Ag]+ at m/z 108.910 were confirmed. Moreover, LCLB56-AgCs exerted an inhibitory effect against all tested bacterial strains (Pseudomonas aeruginosa ATCC10145, Proteus mirabilis ATCC25933, Staphylococcus epidermidis ATCC49461, MSSA ATCC29213, and Staphylococcus aureus ATCC6338). More pronounced antimicrobial effect was noticed for 15 μg/well. Minimum inhibitory concentration required to inhibite the growth of 90% organism (MIC90) of synthetized LCLB56-AgCs was in a range of 3.125–12.5 μg/mL. The concentration at which the viability of the L929 cells was reduced to 50% was above 200 μg/mL for LCLB56-AgNCs. These results open up possibilities for many applications of bioactive silver composites (BioAgCs) synthesized by L. lactis 56 in food and pharmaceutical industries.
... SELDIand MALDI-TOF MS offer great potential for proteome profiling, and have become promising tools for the control of serum quality and assessment of sample handling artifacts. For MS-based protein profiling of clinical samples, commonly used preparation methods are C4/ C18 zip-tip resin [2,15], C8/C18-derivatized magnetic beads [3,[16][17][18][19]21], or anion-exchange chromatography purification [12]. However, these methods are laborand time-intensive due to the labor-intensive procedures involved, including repeated centrifugation steps, multiple steps of sample transfer, washing and elution, and are therefore of limited use in the clinical settings, where sample numbers usually reach several hundreds or even more. ...
Background
High quality clinical samples are critical for meaningful interpretation of data obtained in both basic and translational medicine. More specifically, optimized pre-analysis handling to bio-sample is crucial for avoiding biased analysis in a clinical setting. A universally applicable method for the evaluation of sample quality and pre-analysis handling is therefore in great demand.
Methods
The fingerprint pattern of low molecular weight (LMW) peptides in sera is directly associated with sample quality and handling process. Previous studies for enrichment/isolation of LMW peptides have shown that LMW peptides can be enriched by silica meso-porous material in a sensitive and high-throughput manner. Here, a peptide profile approach utilizing mesoporous silica chip-based sample preparation combined with MALDI MS analysis was used as a new platform for evaluation of bio-sample quality. Rat sera were selected as model sample and analyzed according to their LMW peptide fingerprint spectra.
Results
This novel method can complete the entire sample preparation procedure in a short period of time (<40 min), requires minimum amounts of sample (<10 µL), is of high sensitivity (LOD 10 ng/mL) as well as high reproducibility (CV% < 15%). According to the acquired LMW peptide spectra, we were able to distinguish the serum samples processed under different conditions (including different storage temperature, time, and freezing/thaw cycles) with the help of bioinformatics tools (principle composition analysis and significant difference analysis), and identify the samples that had significantly changed due to the inappropriate processing. Based on the percentage of significantly changed peaks in LMW peptide mass spectrum after handling, a judgment standard was established that can be used to evaluate the status of preservation of a biological sample. In addition, our principle study established recommendations for storage time, storage temperature and freeze/thaw conditions.
Conclusion
Our novel method for analysis of bio-samples allows for effective identification of variations in composition within samples, and provides a cost-effective tool for simple sample manipulation in a clinical setting.
... In MALDI-TOF MS profiling, the major noise source is a chemical noise that originates mainly from the matrix and its cluster-derived signals. Baseline estimation and subtraction are based either on nonparametric heuristic methods or on algorithm approaches using top hat filter [27,89,90,94,96], local regression (LOESS) [91], asymmetric least squares algorithm, convex hull baseline [97,98], statisticssensitive non-linear iterative peak-clipping (SNIP) [99], and waveletbased method [100]. As systematic time shifts may be observed for separate MALDI-TOF MS measurements, recalibration of spectra is necessary. ...
... A list of reference masses can be also used in spectrum quality filtering process and the spectra characterized by low number of reference peaks (below accepted threshold) may be excluded from further processing. The peak list is generally picked using signal-to-noise ratios (S/N) derived from [95] peak area or maximum intensity [90,91,[94][95][96]101,102]. This step is highly affected by prior appropriate estimation of the spectra noise. ...
... The isotopic quality control (QC) value is calculated by comparing the measured isotopic pattern with the theoretical isotopic pattern and relating the result to the noise, as reported previously (Supporting Information, Figure S1). 40 Briefly, the difference between the measured (S i (obs)) pattern and theoretical (S i (est)) pattern is calculated per isotope (i), squared, and divided by the noise (N) squared. The square root of the sum of all isotope QC values yields the analyte QC value (eq 2). ...
... For MALDI-TOF-MS TPNG glycan data, a window of m/z ±20 showed optimal results when comparing m/z ranges of ±50, 45,40,35,30,25,20,15,10,6, and 1 (data not shown). The difference in the optimum is likely due to the difference in sample complexity (TPNG contains far more signals than mAb1 spectra), and an m/z range of 20 for background determination is currently the default value for this definable setting. ...
