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Pyrolysis oil (PO) is an inexpensive biofuel produced via the fast pyrolysis of lignocellulosic biomass. However, PO is unstable and has low energy density, making it unsuitable as a fuel in conven...

Matrix-assisted laser desorption/ionization (MALDI)-MS imaging has been utilized to image a variety of biomolecules, including neuropeptides. Washing a tissue section is an effective way to eliminate interfering background and improve detection of low concentration target analyte molecules; however, many previous methods have not been tested for neuropeptide analysis via MALDI-MS imaging. Using crustacean as a neurological model organism, we developed a new, simple washing procedure and applied this method to characterize neuropeptide changes due to hypoxia stress. By using a 10 second 50:50 EtOH:H(2)O wash, neuropeptide coverage was improved by 1.15-fold, while normalized signal intensities were increased by 5.28-fold.Specifically, hypoxia and hypercapnia stress conditions were investigated due to their environmental relevance to marine invertebrates. Many neuropeptides, including RFamides, pyrokinin, and cardioactive peptides, showed distinct up- and down-regulation for specific neuropeptide isoforms. Since crustacean neuropeptides are homologous to those found in humans, results from these studies can be applied to understand potential roles of neuropeptides involved in medical hypoxia and hypercapnia.

Biomass conversion to alcohols using supercritical methanol depolymerization and hydrodeoxygenation (SCM-DHO) with CuMgAl mixed metal oxide is a promising process for biofuel production. We demonstrate how maple wood can be converted at high weight loadings and product concentrations in a batch and a semi-continuous reactor to a mixture of C2–C10 linear and cyclic alcohols. Maple wood was solubilized semi-continuously in supercritical methanol and then converted to a mixture of C2–C9 alcohols and aromatics over a packed bed of CuMgAlOx catalyst. Up to 95 wt% of maple wood can be solubilized in the methanol by using four temperature holds at 190, 230, 300, and 330 °C. Lignin was solubilized at 190 and 230 °C to a mixture of monomers, dimers, and trimers while hemicellulose and cellulose solubilized at 300 and 330 °C to a mixture of oligomeric sugars and liquefaction products. The hemicellulose, cellulose, and lignin were converted to C2–C10 alcohol fuel precursors over a packed bed of CuMgAlOx catalyst with 70–80% carbon yield of the entire maple wood. The methanol reforming activity of the catalyst decreased by 25% over four beds of biomass, which corresponds to 5 turnovers for the catalyst, but was regenerable after calcination and reduction. In batch reactions, maple wood was converted at 10 wt% in methanol with 93% carbon yield to liquid products. The product concentration can be increased to 20 wt% by partially replacing the methanol with liquid products. The yield of alcohols in the semi-continuous reactor was approximately 30% lower than in batch reactions likely due to degradation of lignin and cellulose during solubilization. These results show that solubilization of whole biomass can be separated from catalytic conversion of the intermediates while still achieving a high yield of products. However, close contact of the catalyst and the biomass during solubilization is critical to achieve the highest yields and concentration of products.

Neuropeptides are low abundance signaling molecules that modulate almost every physiological process, and dysregulation of neuropeptides is implicated in disease pathology. Mass spectrometry (MS) imaging is becoming increasingly useful for studying neuropeptides as new sample preparation methods for improving neuropeptide detection are developed. In particular, proper tissue washes prior to MS imaging have shown to be quick and effective strategies for increasing the number of detectable neuropeptides. Treating tissues with solvents could result in either gain or loss of detection of analytes, and characterization of these wash effects is important for studies targeting sub-classes of neuropeptides. In this communication, we apply aqueous tissue washes that contain sodium phosphate salts, including 10% neutral buffered formalin (NBF), on crustacean brain tissues. Our optimized method resulted in complementary identification of neuropeptides between washed and unwashed tissues, indicating that our wash protocol may be used to increase total neuropeptide identifications. Finally, we show that identical neuropeptides were detected between tissues treated with 10% NBF and an aqueous 1% w/v sodium phosphate solution (composition of 10% NBF without formaldehyde), suggesting that utilizing a salt solution wash affects neuropeptide detection and formaldehyde does not affect neuropeptide detection when our wash protocol is performed.

Mass spectrometry imaging (MSI) is a powerful tool that enables untargeted investigations into the spatial distribution of molecular species in a variety of samples. It has the capability to image thousands of molecules, such as metabolites, lipids, peptides, proteins, and glycans, in a single experiment without labeling. The combination of information gained from mass spectrometry (MS) and visualization of spatial distributions in thin sample sections makes this a valuable chemical analysis tool useful for biological specimen characterization. After minimal but careful sample preparation, the general setup of an MSI experiment involves defining an (x, y) grid over the surface of the sample, with the grid area chosen by the user. The mass spectrometer then ionizes the molecules on the surface of the sample and collects a mass spectrum at each pixel on the section, with the resulting spatial resolution defined by the pixel size. After collecting the spectra, computational software can be used to select an ...

Mass spectrometry-based stable isotope labeling provides the advantages of multiplexing capability and accurate quantification but requires tailored bioinformatics tools for data analysis. Despite the rapid advancements in analytical methodology, it is often challenging to analyze stable isotope labeling-based metabolomics data, particularly for isobaric labeling using MS/MS reporter ions for quantification. We report Metandem, a novel online software tool for isobaric labeling-based metabolomics, freely available at http://metandem.com/web/. Metandem provides a comprehensive data analysis pipeline integrating feature extraction, metabolite quantification, metabolite identification, batch processing of multiple data files, online parameter optimization for custom datasets, data normalization, and statistical analysis. Systematic evaluation of the Metandem tool was demonstrated on UPLC-MS/MS, nanoLC-MS/MS, CE-MS/MS and MALDI-MS platforms, via duplex, 4-plex, 10-plex, and 12-plex isobaric labeling experiments and the application to various biological samples.

We recently developed a novel amine-reactive mass-defect-based chemical tag, dimethyl pyrimidinyl ornithine (DiPyrO), for quantitative proteomic analysis at the MS(1) level. In this work, we further extend the application of the DiPyrO tag, which provides amine group reactivity, optical detection capability, and improved electrospray sensitivity, to quantify N-linked glycans enzymatically released from glycoproteins in the glycosylamine form. Duplex DiPyrO tags that differ in mass by 45.3 mDa were used to label the glycosylamine moieties of freshly released N-glycosylamines from glycoprotein standards and human serum proteins. We demonstrate that both MALDI-LTQ-Orbitrap and nano-HILIC LC/MS/MS Fusion Lumos Orbitrap platforms are capable of resolving the singly or multiply charged N-glycans labeled with mass-defect DiPyrO tags. Dynamic range of quantification, based on MS(1) peak intensities, was evaluated across 2 orders of magnitude. With optimized N-glycan release conditions, glycosylamine labeling conditions, and MS acquisition parameters, the N-glycan profiles and abundances in human serum proteins of cancer patients before and after chemotherapy were compared. Moreover, this study also opens a door for using well-developed amine-reactive tags for relative quantification of glycans, which could be widely applied.

Mass spectrometry-based stable isotope labeling has become a key technology for protein and small-molecule analyses. We developed a multiplexed quantification method for simultaneous proteomics and amine metabolomics analyses via nano reversed-phase liquid chromatography–tandem mass spectrometry (nanoRPLC–MS/MS), called mass defect-based N,N-dimethyl leucine (mdDiLeu) labeling. The duplex mdDiLeu reagents were custom-synthesized with a mass difference of 20.5 mDa, arising from the subtle variation in nuclear binding energy between the two DiLeu isotopologues. Optimal MS resolving powers were determined to be 240K for labeled peptides and 120K for labeled metabolites on the Orbitrap Fusion Lumos instrument. The mdDiLeu labeling does not suffer from precursor interference and dynamic range compression, providing excellent accuracy for MS1-centric quantification. Quantitative information is only revealed at high MS resolution without increasing spectrum complexity and overlapping isotope distribution. Chromatographic performance of polar metabolites was dramatically improved by mdDiLeu labeling with modified hydrophobicity, enhanced ionization efficiency, and picomole levels of detection limits. Paralleled proteomics and amine metabolomics analyses using mdDiLeu were systematically evaluated and then applied to pancreatic cancer cells.