Sequential Extraction of Petroleum Asphaltenes with Magnesium Oxide: A Method To Reduce Complexity and Improve Heteroatom Identity

Sequential extraction of an asphaltene sample with magnesium oxide nanoparticles provides an approach to remove selectively molecules from a complex fraction. This method of extraction relies, in part, on adsorption preferences for differing heteroatoms to leave weakly adsorbing sample constituents in a toluene solution. The extracted sample exhibits reduced complexity enabling more reliable identification of the remaining molecules in solution. Mass spectrometry (MS) data indicate a general bias for preferential removal of higher molecular weight species. This is supported by a shift in the average <i>m</i>/<i>z</i> ratio of the asphaltene distribution to lower <i>m</i>/<i>z</i>, as well as a decrease in the intensity observed for higher <i>m</i>/<i>z</i> ion signals, for higher extraction numbers. UV–vis absorption data corroborate MS data to provide an appreciable visual means to quantify sample uptake after several sequential extraction steps with MgO. Furthermore, both UV and MS data indicate a point of diminishing returns, after which subsequent extraction with MgO nanoparticles results in limited adsorption of remaining asphaltene constituents. The remaining asphaltene constituents can then be treated with NiO nanoparticles in order to identify molecules containing pyridyl functional groups. Implementation of a more exhaustive MgO extraction, prior to treatment with NiO, resulted in an improved method for profiling pyridyl-containing structures in a complex asphaltene mixture relative to previous work

Exploiting Metal Oxide Nanoparticle Selectivity in Asphaltenes for Identification of Pyridyl-Containing Molecules

Extraction efficiencies for a series of model compounds representing heteroatom functional groups believed to be present in asphaltenes were determined in batch extractions with a variety of metal oxide nanoparticles (Fe₃O₄, TiO₂, NiO, Co₃O₄, and MgO). Extraction efficiencies from toluene solution varied depending upon both the adsorbate and the type of metal oxide used for extraction. However, the adsorbate was found to be the most important factor governing selectivity, which generally followed the trend: benzoic acid ≫ pyridine ≈ phenol > pyrrole > thiophene ≈ diphenylsulfide ≈ benzophenone. An important exception to this trend was that MgO did not appreciably adsorb pyridine. The divergent adsorption behavior of pyridine on NiO (extraction efficiency = 82 ± 1%) and MgO (extraction efficiency = 0 ± 2%) was subsequently exploited to demonstrate a novel approach for profiling pyridine-containing molecules in an authentic asphaltene sample. Specifically, mass spectra of the asphaltene mixture were obtained before and after treatment with NiO or MgO and compared to identify peaks exhibiting reduced intensity after treatment with NiO but no appreciable change in intensity after treatment with MgO. Results of batch extraction studies with model compounds and elemental composition data deduced from accurate mass measurements support that these peaks likely correspond to (or minimally contain) a molecule(s) possessing a pyridyl functional group

Extending Proteome Coverage by Combining MS/MS Methods and a Modified Bioinformatics Platform Adapted for Database Searching of Positive and Negative Polarity 193 nm Ultraviolet Photodissociation Mass Spectra

To extend proteome coverage obtained from bottom-up mass spectrometry approaches, three complementary ion activation methods, higher energy collision dissociation (HCD), ultraviolet photodissociation (UVPD), and negative mode UVPD (NUVPD), are used to interrogate the tryptic peptides in a human hepatocyte lysate using a high performance Orbitrap mass spectrometer. The utility of combining results from multiple activation techniques (HCD+UVPD+NUVPD) is analyzed for total depth and breadth of proteome coverage. This study also benchmarks a new version of the Byonic algorithm, which has been customized for database searches of UVPD and NUVPD data. Searches utilizing the customized algorithm resulted in over 50% more peptide identifications for UVPD and NUVPD tryptic peptide data sets compared to other search algorithms. Inclusion of UVPD and NUVPD spectra resulted in over 600 additional protein identifications relative to HCD alone

Extending Proteome Coverage by Combining MS/MS Methods and a Modified Bioinformatics Platform Adapted for Database Searching of Positive and Negative Polarity 193 nm Ultraviolet Photodissociation Mass Spectra

To extend proteome coverage obtained from bottom-up mass spectrometry approaches, three complementary ion activation methods, higher energy collision dissociation (HCD), ultraviolet photodissociation (UVPD), and negative mode UVPD (NUVPD), are used to interrogate the tryptic peptides in a human hepatocyte lysate using a high performance Orbitrap mass spectrometer. The utility of combining results from multiple activation techniques (HCD+UVPD+NUVPD) is analyzed for total depth and breadth of proteome coverage. This study also benchmarks a new version of the Byonic algorithm, which has been customized for database searches of UVPD and NUVPD data. Searches utilizing the customized algorithm resulted in over 50% more peptide identifications for UVPD and NUVPD tryptic peptide data sets compared to other search algorithms. Inclusion of UVPD and NUVPD spectra resulted in over 600 additional protein identifications relative to HCD alone

Extending Proteome Coverage by Combining MS/MS Methods and a Modified Bioinformatics Platform Adapted for Database Searching of Positive and Negative Polarity 193 nm Ultraviolet Photodissociation Mass Spectra

To extend proteome coverage obtained from bottom-up mass spectrometry approaches, three complementary ion activation methods, higher energy collision dissociation (HCD), ultraviolet photodissociation (UVPD), and negative mode UVPD (NUVPD), are used to interrogate the tryptic peptides in a human hepatocyte lysate using a high performance Orbitrap mass spectrometer. The utility of combining results from multiple activation techniques (HCD+UVPD+NUVPD) is analyzed for total depth and breadth of proteome coverage. This study also benchmarks a new version of the Byonic algorithm, which has been customized for database searches of UVPD and NUVPD data. Searches utilizing the customized algorithm resulted in over 50% more peptide identifications for UVPD and NUVPD tryptic peptide data sets compared to other search algorithms. Inclusion of UVPD and NUVPD spectra resulted in over 600 additional protein identifications relative to HCD alone

Extending Proteome Coverage by Combining MS/MS Methods and a Modified Bioinformatics Platform Adapted for Database Searching of Positive and Negative Polarity 193 nm Ultraviolet Photodissociation Mass Spectra

To extend proteome coverage obtained from bottom-up mass spectrometry approaches, three complementary ion activation methods, higher energy collision dissociation (HCD), ultraviolet photodissociation (UVPD), and negative mode UVPD (NUVPD), are used to interrogate the tryptic peptides in a human hepatocyte lysate using a high performance Orbitrap mass spectrometer. The utility of combining results from multiple activation techniques (HCD+UVPD+NUVPD) is analyzed for total depth and breadth of proteome coverage. This study also benchmarks a new version of the Byonic algorithm, which has been customized for database searches of UVPD and NUVPD data. Searches utilizing the customized algorithm resulted in over 50% more peptide identifications for UVPD and NUVPD tryptic peptide data sets compared to other search algorithms. Inclusion of UVPD and NUVPD spectra resulted in over 600 additional protein identifications relative to HCD alone

Extending Proteome Coverage by Combining MS/MS Methods and a Modified Bioinformatics Platform Adapted for Database Searching of Positive and Negative Polarity 193 nm Ultraviolet Photodissociation Mass Spectra

To extend proteome coverage obtained from bottom-up mass spectrometry approaches, three complementary ion activation methods, higher energy collision dissociation (HCD), ultraviolet photodissociation (UVPD), and negative mode UVPD (NUVPD), are used to interrogate the tryptic peptides in a human hepatocyte lysate using a high performance Orbitrap mass spectrometer. The utility of combining results from multiple activation techniques (HCD+UVPD+NUVPD) is analyzed for total depth and breadth of proteome coverage. This study also benchmarks a new version of the Byonic algorithm, which has been customized for database searches of UVPD and NUVPD data. Searches utilizing the customized algorithm resulted in over 50% more peptide identifications for UVPD and NUVPD tryptic peptide data sets compared to other search algorithms. Inclusion of UVPD and NUVPD spectra resulted in over 600 additional protein identifications relative to HCD alone

Characterization of Slow-Pyrolysis Bio-Oils by High-Resolution Mass Spectrometry and Ion Mobility Spectrometry

Bio-oils produced from biomass pyrolysis are an attractive fuel source that requires significant upgrading. Before upgrade strategies can be developed, the molecular composition of bio-oils needs to be better understood. In this work, oily and aqueous fractions of bio-oils produced by slow pyrolysis of two feedstocks, pine shavings (PS) and corn stover (CS), were analyzed by negative electrospray ionization (ESI)-Orbitrap and ion mobility-time-of-flight mass spectrometry (IM-TOF-MS). Analyte ion signal was observed primarily between <i>m</i>/<i>z</i> 80 and 450 in the mass spectra of these samples. Mass defect analysis and collision-induced dissociation (CID) experiments performed on mobility-separated ions indicated a high degree of homology among bio-oil samples produced from both feedstocks. Oxygen-rich species having between 1 and 9 oxygen atoms and with double bond equivalents (DBEs) ranging from 1 to 15 were identified, indicating that catalytic upgrading will likely be required if slow-pyrolysis bio-oils are to be utilized as fuel. IM-MS and IM-MS/MS analysis of ions belonging to select CH₂-homologous series suggest that mass-mobility correlations and post-ion mobility CID mass spectra may be useful in defining structural relationships among members of a given Kendrick mass defect series

Parsimonious Charge Deconvolution for Native Mass Spectrometry

Charge deconvolution infers the mass from mass over charge (<i>m</i>/<i>z</i>) measurements in electrospray ionization mass spectra. When applied over a wide input <i>m</i>/<i>z</i> or broad target mass range, charge-deconvolution algorithms can produce artifacts, such as false masses at one-half or one-third of the correct mass. Indeed, a maximum entropy term in the objective function of MaxEnt, the most commonly used charge deconvolution algorithm, favors a deconvolved spectrum with many peaks over one with fewer peaks. Here we describe a new “parsimonious” charge deconvolution algorithm that produces fewer artifacts. The algorithm is especially well-suited to high-resolution native mass spectrometry of intact glycoproteins and protein complexes. Deconvolution of native mass spectra poses special challenges due to salt and small molecule adducts, multimers, wide mass ranges, and fewer and lower charge states. We demonstrate the performance of the new deconvolution algorithm on a range of samples. On the heavily glycosylated plasma properdin glycoprotein, the new algorithm could deconvolve monomer and dimer simultaneously and, when focused on the <i>m</i>/<i>z</i> range of the monomer, gave accurate and interpretable masses for glycoforms that had previously been analyzed manually using <i>m</i>/<i>z</i> peaks rather than deconvolved masses. On therapeutic antibodies, the new algorithm facilitated the analysis of extensions, truncations, and Fab glycosylation. The algorithm facilitates the use of native mass spectrometry for the qualitative and quantitative analysis of protein and protein assemblies

Extended O‑GlcNAc on HLA Class-I-Bound Peptides

We report unexpected mass spectrometric observations of glycosylated human leukocyte antigen (HLA) class I-bound peptides. Complemented by molecular modeling, <i>in vitro</i> enzymatic assays, and oxonium ion patterns, we propose that the observed O-linked glycans carrying up to five monosaccharides are extended O-GlcNAc’s rather than GalNAc-initiated O-glycans. A cytosolic O-GlcNAc modification is normally terminal and does not extend to produce a polysaccharide, but O-GlcNAc on an HLA peptide presents a special case because the loaded HLA class I complex traffics through the endoplasmic reticulum and Golgi apparatus on its way to the cell membrane and is hence exposed to glycosyltransferases. We also report for the first time natural HLA class I presentation of O- and N-linked glycopeptides derived from membrane proteins. HLA class I peptides with centrally located oligosaccharides have been shown to be immunogenic and may thus be important targets for immune surveillance