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Basics Of Gas Chromatography Mass Spectrometry System
Published in Raquel Cumeras, Xavier Correig, Volatile organic compound analysis in biomedical diagnosis applications, 2018
William Hon Kit Cheung, Raquel Cumeras
Finally, a second stage ion optics focus and guides the ions from the C-trap into orbitrap mass analyzer, as the ions enter the mass analyzer, an increasing voltage is applied to the central spindle electrode. This induces the initial packet of ions to be squeezed towards the electrodes and osculated around it in an orbital motion, as the voltage to the central electrode increases, the radius of ion trajectory decreases due to an effect know as electrodynamics squeezing. The applied voltages ramp is then ceased, and the ion osculation is stabilized and trapped around the central spindle electrode. Ions of different m/z will osculate at independent orbital trajectories with respect to other. As the ions osculate back and forth around the central electrodes, a sinusoidal signal is produced and detected using a differential amplifier. For multiple signals, the super positioning of multiple sinusoidal signals are observed, the different frequencies of the sinusoidal signals equate to the different m/z and are inferred using inverse FT signal transformation.
The role of microtubules in the regulation of epithelial junctions
Published in Tissue Barriers, 2018
Ekaterina Vasileva, Sandra Citi
MgcRacGAP, together with the kinesin-6-family member MKLP1 (mitotic kinesin-like protein), forms the centralspindlin complex, which organizes antiparallel arrays of MTs at the spindle midzone and midbody, controls the activity of RhoA and Rac1 during furrow ingression, and links the mitotic spindle to the plasma membrane during cytokinesis.129 ECT2 is a Rho GEF, which concentrates on the central spindle by binding to centralspindlin, and is essential for RhoA activation in the cleavage furrow.130,131 MgcRacGAP localizes to MT plus-ends at the equatorial cortex as cytokinesis initiates, and it tracks with EB3 at growing MT plus-ends.132 In contrast, in interphase confluent epithelial cells, MgcRacGAP accumulates at apical junctions, where it is recruited by cingulin, paracingulin and α-catenin133,134 (Figure 3). In some but not all cell types the centralspindlin complex recruits the ECT2 to the ZA133,134 (Figure 3). MgcRacGAP regulates AJ but not TJ structure, through its function as a GTPase activating protein (GAP) activity, and signaling via the RhoA pathway.135 The C-terminal region of MgcRacGAP is involved to its targeting both to AJ, via a SxIP motif,132 and to TJ, via its interaction with cingulin and paracingulin.134 These observations suggest that initially MgcRacGAP is delivered to AJ through plus-end directed motors, and then is redistributed at both TJ and AJ through multiple interactions.
Liquid chromatography coupled to mass spectrometry for metabolite profiling in the field of drug discovery
Published in Expert Opinion on Drug Discovery, 2019
Javier Saurina, Sonia Sentellas
Orbitrap analyzers are one of the newest developed HRMS instruments. The resolution principle relies on ions trapped in the orbitrap core cell where the central spindle-like and the outer electrodes create a special geometry electrostatic field [75]. The ions undergo complex oscillations rotating around the central electrode but also axially moving along its length. High-resolution spectra are then obtained measuring the intensity and frequency of the oscillating ions (transients). A more comprehensive description of orbitrap technology and acquisition modes can be found elsewhere [75,76]. The spectral resolution from these systems is much higher than that obtained with TOF instruments. However, as the spectral resolution directly depends on the scan time, when dealing with narrow chromatographic peaks such as those from UHPLC, in practice, resolutions attained with both orbitrap and TOF analyzers are similar. Different processing and filtering strategies can be followed to face metabolite identification. Rashid and coworkers identified the two main metabolites of PP242, an ATP-competitive selective dual inhibitor of mTOR, using a linear ion trap-Orbitrap MS platform (LTQ-Orbitrap). Mass differences between the metabolites and the parent compound and MS/MS fragmentation patterns, together with the fundamental knowledge of xenobiotic biotransformation, were key for the tentative structure assignation [82]. Here, again, post-acquisition processing methods are fundamental when tens of metabolites are simultaneously generated. For instance, the metabolism of 6ʹ-hydroxy-3,4,5,2ʹ,4ʹ-pentamethoxychalcone, considered to be the major active ingredient (with anti-allergic, anti-oxidant, anti-bacterial, anti-proliferative and anti-inflammatory activities) of Murraya paniculata, was studied by Liu et al. [83]. A total of 60 and 35 metabolites were identified in rat urine and feces, respectively based on data mining techniques including high-resolution extracted ion chromatogram (HREIC) and MMDF. HREIC was especially useful to detect and confirm those expected metabolites (i.e. those with m/z values easily predicted from the parent drug structure). On the other hand, for unpredicted (fully unknown) metabolites, well-defined MDF templates (drug filter, substructure filter and conjugate filter) were mandatory for a tentative identification of compounds. Other similar applications can be found elsewhere for dealing with puerarin and epicatechin metabolites in rats [84,85].