Comparison of Compound Identification Tools Using Data Dependent and Data Independent High-Resolution Mass Spectrometry Spectra.

Liquid chromatography combined with high-resolution mass spectrometry (LC-HRMS) is a frequently applied technique for suspect screening (SS) and non-target screening (NTS) in metabolomics and environmental toxicology. However, correctly identifying compounds based on SS or NTS approaches remains challenging, especially when using data-independent acquisition (DIA). This study assessed the performance of four HRMS-spectra identification tools to annotate in-house generated data-dependent acquisition (DDA) and DIA HRMS spectra of 32 pesticides, veterinary drugs, and their metabolites. The identification tools were challenged with a diversity of compounds, including isomeric compounds. The identification power was evaluated in solvent standards and spiked feed extract. In DDA spectra, the mass spectral library mzCloud provided the highest success rate, with 84% and 88% of the compounds correctly identified in the top three in solvent standard and spiked feed extract, respectively. The in silico tools MSfinder, CFM-ID, and Chemdistiller also performed well in DDA data, with identification success rates above 75% for both solvent standard and spiked feed extract. MSfinder provided the highest identification success rates using DIA spectra with 72% and 75% (solvent standard and spiked feed extract, respectively), and CFM-ID performed almost similarly in solvent standard and slightly less in spiked feed extract (72% and 63%). The identification success rates for Chemdistiller (66% and 38%) and mzCloud (66% and 31%) were lower, especially in spiked feed extract. The difference in success rates between DDA and DIA is most likely caused by the higher complexity of the DIA spectra, making direct spectral matching more complex. However, this study demonstrates that DIA spectra can be used for compound annotation in certain software tools, although the success rate is lower than for DDA spectra.

Acute cytomegalovirus infection modulates the intestinal microbiota and targets intestinal epithelial cells.

Primary and recurrent cytomegalovirus (CMV) infections frequently cause CMV colitis in immunocompromised as well as inflammatory bowel disease (IBD) patients. Additionally, colitis occasionally occurs upon primary CMV infection in patients who are apparently immunocompetent. In both cases, the underlying pathophysiologic mechanisms are largely elusive - in part due to the lack of adequate access to specimens. We employed the mouse cytomegalovirus (MCMV) model to assess the association between CMV and colitis. During acute primary MCMV infection of immunocompetent mice, the gut microbial composition was affected as manifested by an altered ratio of the Firmicutes to Bacteroidetes phyla. Interestingly, these microbial changes coincided with high-titer MCMV replication in the colon, crypt hyperplasia, increased colonic pro-inflammatory cytokine levels, and a transient increase in the expression of the antimicrobial protein Regenerating islet-derived protein 3 gamma (Reg3γ). Further analyses revealed that murine and human intestinal epithelial cell lines, as well as primary intestinal crypt cells and organoids represent direct targets of CMV infection causing increased cell death. Accordingly, in vivo MCMV infection disrupted the intestinal epithelial barrier and increased apoptosis of intestinal epithelial cells. In summary, our data show that CMV transiently induces colitis in immunocompetent hosts by altering the intestinal homeostasis.

A strategy to determine the fate of active chemical compounds in soil; applied to antimicrobially active substances.

Data on the fate of chemical substances in the environment after e.g. manure application is mandatory input for risk assessment in perspective of a more circular biobased economy. Such fate studies include a persistence study to determine a half-life value and a mobility study. It is recognized that not only the native substance should be considered, but that also degradation products should be included that might exert a similar effect as the native substance. We report a tiered fate study strategy that starts with a persistence study. For non-persistent substances a study is performed to determine if degradation products have a similar effect as the native compound. If so, a procedure using high resolution mass spectrometry is suggested to identify the potentially active degradation products. Based on the outcomes, substances are divided into three categories: (I) persistent, (II) degradable to inactive products or (III) degradable to active products. Even though the priority is with category I and III, for all substances and possible degradation products a mobility study is proposed. The fate strategy is successfully applied to ten antimicrobially active substances originating from the tetracyclines, sulfonamides, diaminopyrimidines, fluoroquinolones, macrolides and lincosamides. The fluoroquinolones, tetracyclines and trimethoprim were relatively persistent. The sulfonamides, macrolides and lincomycin (the latter also depending on soil type) degraded relatively quickly. Tylosin A proved to degrade to antimicrobially active degradation products which were tentitatively identified as tylosin C, tylosin A acid, tylosin B acid and tylosin C acid.

Adduct ion formation as a tool for the molecular structure assessment of ten isomers in traveling wave and trapped ion mobility spectrometry.

RATIONALE: The separation of isomeric compounds with major differences in their physiochemical and pharmacokinetic properties is of particular importance in pharmaceutical R&D. However, the structural assessment and separation of these compounds with current analytical techniques and methods are still a challenge. In this study, we describe strategies to separate the various structural and stereo-isomers. METHODS: The separation of ten structural and stereo-isomers was investigated using Trapped and Travelling Wave ion mobility spectrometry (TIMS and TWIMS). Different strategies including adduct ion formation with Na, Li, Ag and Cs as well as fragmentation before and after the ion mobility cell were applied to separate the isomeric compounds. RESULTS: All the counter ions (in particular Na) strongly coordinated with the test analytes in all the IMS systems. The highest resolving power was achieved for the sodium and lithium adducts using TIMS-time-of-flight (TOF). However, some separation was attained on a Synapt HDMS system with its unique potential to monitor the ion mobility of the product ions. The elution order of the adduct ions was the same in all instruments, in which, unexpectedly, the para-substituted isomer of the [M + Na]+ species had the lowest collision cross section followed by the meta- and ortho-isomers. CONCLUSIONS: The formation of adduct ions could facilitate the separation of structural and even stereo-isomers by generating different molecular conformations. In addition, fragmenting isomers before or after the ion mobility cell is a valuable strategy to separate and also to assess the structures of adducts and different conformers.

Comparison of structure and transport properties of concentrated hard and soft sphere fluids.

Using Newtonian and Brownian dynamics simulations, the structural and transport properties of hard and soft spheres have been studied. The soft spheres were modeled using inverse power potentials (V approximately r(-n), with 1n the potential softness). Although, at constant density, the pressure, diffusion coefficient, and viscosity depend on the particle softness up to extremely high values of n, we show that scaling the density with the freezing point for every system effectively collapses these parameters for n > or = 18 (including hard spheres) for large densities. At the freezing points, the long range structure of all systems is identical, when length is measured in units of the interparticle distance, but differences appear at short distances (due to the different shapes of the interaction potential). This translates into differences at short times in the velocity and stress autocorrelation functions, although they concur to give the same value of the corresponding transport coefficient (for the same density to freezing ratio); the microscopic dynamics also affects the short time behavior of the correlation functions and absolute values of the transport coefficients, but the same scaling with the freezing density works for Newtonian or Brownian dynamics. For hard spheres, the short time behavior of the stress autocorrelation function has been studied in detail, confirming quantitatively the theoretical forms derived for it.