Journal: Molecular & Cellular Proteomics : MCP

Article Title: Advancing a High Throughput Glycotope-centric Glycomics Workflow Based on nanoLC-MS2-product Dependent-MS3 Analysis of Permethylated Glycans* *

doi: 10.1074/mcp.TIR117.000156

Figure Lengend Snippet: RP C18 nanoLC separation of permethylated high mannose type and biantennary complex type N-glycans. Overlay plots of extracted ion chromatograms for the high mannose structures indicated that the Man 5–9 GlcNAc 2 itol N-glycans were well resolved from one another but not for their individual isomeric constituents. Larger complex type N-glycans, with and without sulfates, produced more complicated and not well resolved ion chromatograms, the unambiguous identification of individual chromatographic peaks based on MS 2 of the protonated molecular ions alone is not feasible but glycotopes carried on each can still be efficiently determined by MS 2 -pd-MS 3 analysis.

Article Snippet: Another EASY-nLC™ 1200 system was interfaced to an Orbitrap Fusion™ Lumos™ Tribrid™ Mass Spectrometer (ThermoFisher Scientific) via a Nanospray Flex™ Ion Sources (ThermoFisher Scientific) for nanoLC separation at 50 °C using the same 25 cm x 75 μm C18 column (Acclaim PepMap® RSLC, ThermoFisher Scientific) at a constant flow rate of 300 nL/min.

Techniques: Produced, Mass Spectrometry

Journal: Molecular & Cellular Proteomics : MCP

Article Title: Advancing a High Throughput Glycotope-centric Glycomics Workflow Based on nanoLC-MS2-product Dependent-MS3 Analysis of Permethylated Glycans* *

doi: 10.1074/mcp.TIR117.000156

Figure Lengend Snippet: RP C18 nanoLC separation and MS 2 /MS 3 identification of permethylated, nonfucosylated and mono-fucosylated, single LacNAc-extended core 1 and core 2 O-glycan structures. Overlay plots of extracted ion chromatograms for the nonfucosylated ( m / z 961.53) and monofucosylated ( m / z 1135.62) (Hex-HexNAc)-Hex-HexNAcitol derived from AGC and Colo205 cells are shown to demonstrate ability to resolve the smaller isomeric O-glycans. Peaks are labeled #1 - 9, the MS 2 and target MS 3 (small inset) spectra of which are shown in 9 small panels, with the diagnostic oxonium ions annotated in red. Z 1 ion for the reducing end GalNAitol (annotated in blue) could be readily detected at m / z 294 or after further loss of 6-arm substituents at m / z 280, which is informative of core 1 versus core 2 structures. A Z 2 ion at m / z 498 is also commonly observed for all core structures that are extended at either the 6 or 3-arm but not both. Extended core 1 structure further afforded a B ion at the Gal of the Gal-GalNAcitol, e.g. m / z 668 and 842, which is very useful to define the entire moiety extending from the 3-arm. Under low collision energy, protonated permethylated glycans do not normally yield cross-ring cleavage ions.

Article Snippet: Another EASY-nLC™ 1200 system was interfaced to an Orbitrap Fusion™ Lumos™ Tribrid™ Mass Spectrometer (ThermoFisher Scientific) via a Nanospray Flex™ Ion Sources (ThermoFisher Scientific) for nanoLC separation at 50 °C using the same 25 cm x 75 μm C18 column (Acclaim PepMap® RSLC, ThermoFisher Scientific) at a constant flow rate of 300 nL/min.

Techniques: Mass Spectrometry, Derivative Assay, Labeling, Diagnostic Assay

Journal: Molecular & Cellular Proteomics : MCP

Article Title:

doi: 10.1074/mcp.O114.046573

Figure Lengend Snippet: Histone preparation with Lys-propionylation, trypsin digestion, and N-terminal PIC-labeling. A , Sample preparation workflow. B , Relative ionization efficiencies H3 T3-R8 peptide in all modified forms. C , Recovery of H3 T3-R8 peptides following StageTip C18 cleanup. All abundances are expressed relative to the unmodified peptide.

Article Snippet: Peptides were loaded onto a C18 column (BEH-C18; 100 μm i.d. x 10 cm; 1.7 μm particles, 130Å pores; Waters Corp., Milford, MA) for 10 min at 1.5 μl per minute in 2% solvent B (0.1% v/v formic acid, 98% v/v acetonitrile) and separated at 1 μl per minute by a linear gradient from 2% solvent B to 25% solvent B over 60 min followed by a ramp to 40% B in 15 min, then to 90% B and re-equilibration at 2% B for a 90-min total run time.

Techniques: Labeling, Sample Prep, Modification

Journal: Molecular & Cellular Proteomics : MCP

Article Title:

doi: 10.1074/mcp.O114.046573

Figure Lengend Snippet: Sources of discrepancy in H3K4 quantitation. A , Equimolar mixture of isotope-labeled, propionylated H3 T3-R8 peptide in eight modification states, injected in the milieu of endogenous histones from 293T cells. B , Time course of digestion releasing propionylated H3 T3-R8 from propionylated H3 A1-R17, where K4 is trimethylated. C , Relative ionization efficiencies of the propionylated H3 T3-R8 peptide in its various modified forms. D , Recovery of H3 T3-R8 peptides following StageTip C18 cleanup. In panels A , C , D , all abundances are expressed relative to the unmodified peptide.

Article Snippet: Peptides were loaded onto a C18 column (BEH-C18; 100 μm i.d. x 10 cm; 1.7 μm particles, 130Å pores; Waters Corp., Milford, MA) for 10 min at 1.5 μl per minute in 2% solvent B (0.1% v/v formic acid, 98% v/v acetonitrile) and separated at 1 μl per minute by a linear gradient from 2% solvent B to 25% solvent B over 60 min followed by a ramp to 40% B in 15 min, then to 90% B and re-equilibration at 2% B for a 90-min total run time.

Techniques: Quantitation Assay, Labeling, Modification, Injection

Journal: Molecular & Cellular Proteomics : MCP

Article Title:

doi: 10.1074/mcp.O114.046573

Figure Lengend Snippet: Direct comparison of standard propionylation (Prop-x2) labeling and the hybrid (Prop-PIC) labeling method via LC-MS using a 1:1 mixture of labeled histone samples. A , Extracted ion chromatograms of the hydrophilic H3T3-R8 peptide (TK 4 QTAR) and its modified versions on C18 reverse-phase LC-MS. Retention times and relative abundances are increased by the hybrid labeling method. B , Quantitative comparison of the recoveries of each H3.1 histone tail peptide, where integrated peak areas for all modification states are combined. C , Quantitative comparison of the recoveries of the H3 T3-R8 peptide in its various modification states for the Prop-PIC versus Propx2 (integrated peak areas; n = 3, error bars are standard deviations).

Article Snippet: Peptides were loaded onto a C18 column (BEH-C18; 100 μm i.d. x 10 cm; 1.7 μm particles, 130Å pores; Waters Corp., Milford, MA) for 10 min at 1.5 μl per minute in 2% solvent B (0.1% v/v formic acid, 98% v/v acetonitrile) and separated at 1 μl per minute by a linear gradient from 2% solvent B to 25% solvent B over 60 min followed by a ramp to 40% B in 15 min, then to 90% B and re-equilibration at 2% B for a 90-min total run time.

Techniques: Labeling, Liquid Chromatography with Mass Spectroscopy, Modification

Journal: Scientia Pharmaceutica

Article Title: Identification of Degradation Products and a Stability-Indicating RP-HPLC Method for the Determination of Flupirtine Maleate in Pharmaceutical Dosage Forms

doi: 10.3797/scipharm.1310-01

Figure Lengend Snippet: Optimized chromatogram of flupirtine maleate (10.3 min) on a C18 column

Article Snippet: Method Development and Optimization of the Chromatographic Conditions In preliminary experiments, the drug was subjected to the reversed-phase mode using a C18 column (Agilent, 250 × 4.6 mm, 5μ) and mobile phases consisting of water (pH 3.0 adjusted with orthophosphoric acid) and methanol by varying the % aqueous phase from 10% to 30%.

Techniques:

Journal: BMC Chemical Biology

Article Title: High toxicity and specificity of the saponin 3-GlcA-28-AraRhaxyl-medicagenate, from Medicago truncatula seeds, for Sitophilus oryzae

doi: 10.1186/1472-6769-12-3

Figure Lengend Snippet: Chromatograms of Medicago truncatula seed extract. A. Injection into a C18 column (250 × 25 mm, 5 μm, Phenomenex) on an Agilent 1200 HPLC apparatus. The flow was 3.5 mL.min -1 . The gradient was H 2 O + 0.04%TFA (solvent A) / ACN +0.04% TFA (solvent B) 90/10 for 5 min, then 60% solvent B for 25 min. The elution was monitored using a diode array detector at 210 nm. B. The fraction purified in A was injected into the same column with an elution under isocratic conditions (solvent B 30%). Peaks containing entomotoxic activity are indicated.

Article Snippet: The extract was injected into a C18 column (250 x 4.1 mm, 3 μm, Phenomenex).

Techniques: Injection, High Performance Liquid Chromatography, Flow Cytometry, Purification, Activity Assay

Journal: Proteomics

Article Title: Using iRT, a normalized retention time for more targeted measurement of peptides

doi: 10.1002/pmic.201100463

Figure Lengend Snippet: Selection of iRT-peptides and definition of iRT-C18 scale

Article Snippet: 100fmol each in 1 μg total protein) and measured in LC-MRM with a 90 min linear gradient (5%–35% acetonitrile as organic modifier with 0.1% formic acid) on a C18 column (Magic C18 AQ resin 3μm particle size/300Å pore size; Michrom, Leonberg, Germany).

Techniques: Selection

Journal: The Journal of Venomous Animals and Toxins Including Tropical Diseases

Article Title: Partial purification and functional characterization of Ts19 Frag-I, a novel toxin from Tityus serrulatus scorpion venom

doi: 10.1186/s40409-015-0051-6

Figure Lengend Snippet: Rechromatography of the subfractions eluted from the fractions VIIIA and VIIIB. ( a ) Subfraction 7. ( b ) Subfraction 9. The C18 column (2.1 mm × 250.0 mm, 3.6 μm particles, Phenomenex) was equilibrated with 0.1 % TFA and the proteins were eluted using a concentration gradient from 0 to 100 % of solution B (80 % ACN in 0.1 % TFA), represented by the dashed line. Absorbance was monitored at 214 nm, at 25 °C, using a FPLC Äkta Purifier UPC-10 system. Fractions of 0.3 mL/tube were collected at a flow rate of 0.4 mL/min

Article Snippet: The fractions VIIIA and VIIIB (4 mg) were submitted to reversed-phase chromatography using a 4.6 mm × 250.0 mm C18 column (5 μm particles, Shimadzu Corp., Japan); the eluted subfractions were rechromatographed on a 2.1 mm × 250.0 mm C18 column (3.6 μm particles, Phenomenex, USA).

Techniques: Concentration Assay, Fast Protein Liquid Chromatography, Flow Cytometry

Journal: The Journal of Venomous Animals and Toxins Including Tropical Diseases

Article Title: Partial purification and functional characterization of Ts19 Frag-I, a novel toxin from Tityus serrulatus scorpion venom

doi: 10.1186/s40409-015-0051-6

Figure Lengend Snippet: Chromatographic profiles of fractions VIIIA and VIIIB from Tsv. ( a ) Fraction VIIIA. ( b ) Fraction VIIIB. Fractions (4 mg, eluted of the cation exchange chromatography from Tityus serrulatus venom) were submitted to RP-FPLC on a C18 column (4.6 mm × 250.0 mm, 5 μm particles, Shimadzu Corp.). The column was equilibrated with 0.1 % trifluoroacetic acid (TFA) and the proteins were eluted using a concentration gradient from 0 to 100 % of solution B (80 % acetonitrile (ACN) in 0.1 % TFA), represented by the dashed line. Absorbance was monitored at 214 nm, at 25 °C, using an FPLC Äkta Purifier UPC-10 system. Fractions of 0.3 mL/tube were collected at a flow rate of 0.7 mL/min

Article Snippet: The fractions VIIIA and VIIIB (4 mg) were submitted to reversed-phase chromatography using a 4.6 mm × 250.0 mm C18 column (5 μm particles, Shimadzu Corp., Japan); the eluted subfractions were rechromatographed on a 2.1 mm × 250.0 mm C18 column (3.6 μm particles, Phenomenex, USA).

Techniques: Chromatography, Fast Protein Liquid Chromatography, Concentration Assay, Flow Cytometry