Glycosyl composition analysis was performed by combined gas chromatography/mass spectrometry (GC/MS) of the per-O-trimethylsilyl (TMS) derivatives of the monosaccharide methyl glycosides produced from the sample by acidic methanolysis as described previously by Santander et al.68 (link). Briefly, the samples (230 and 240 μg) were heated with methanolic HCl in a sealed screw-top glass test tube for 17 h at 80 °C. After cooling and removal of the solvent under a stream of nitrogen, the samples were treated with a mixture of methanol, pyridine, and acetic anhydride for 30 min to re–N-acetylate the hexosamines. The solvents were evaporated, and the samples were derivatized with Tri-Sil (Pierce) at 80 °C for 30 min. GC/MS analysis of the TMS methyl glycosides was performed on an Agilent 7890A GC interfaced to a 5975C MSD, using a Supelco Equity-1 fused silica capillary column (30 m 0.25 mm ID).
Equity 1 fused silica capillary column
Manufactured by Merck Group
Sourced in United States
The Equity-1 fused silica capillary column is a chromatographic column used for the separation and analysis of various chemical compounds. It is made of fused silica, a durable and inert material, and is designed to provide efficient and reliable separation performance. The column's core function is to facilitate the separation of complex mixtures by leveraging the differences in the interactions between the analytes and the stationary phase within the column.
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8 protocols using equity 1 fused silica capillary column
1
Glycosyl Composition Analysis by GC/MS
2
Glycosyl Composition Analysis by GC-MS
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Glycosyl composition analysis was performed by combined gas chromatography—mass spectrometry (GC-MS) of the per-O-trimethylsilyl (TMS) derivatives of the monosaccharide methyl glycosides produced from the sample by acidic methanolysis as described previously by Santander et al. (2013 (link)). Briefly, the sample (500 μg) with myo-inositol (internal standard, 20 μg) was heated with methanolic HCl in a sealed screw-top glass test tube for 18 h at 80°C. After cooling and removal of the solvent under a stream of nitrogen, the samples were treated with a mixture of methanol, pyridine, and acetic anhydride for 30 min. The solvents were evaporated, and the samples were derivatized with Tri-Sil® (Pierce) at 80°C for 30 min. GC-MS analysis of the TMS methyl glycosides was performed on an Agilent 7890A GC interfaced to a 5975C MSD, using an Supelco Equity-1 fused silica capillary column (30 m × 0.25 mm ID).
3
Glycosyl Composition Analysis by GC/MS
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Glycosyl composition analysis was performed by combined gas chromatography/mass spectrometry (GC/MS) of the per-O-trimethylsilyl (TMS) derivatives of the monosaccharide methyl glycosides produced from the sample by acidic methanolysis as described previously (36 (link)). An aliquot equal to 0.3 mg sample weight was pipetted from each sample solution. To this, 20 µg inositol was added as internal standard. The solution was then lyophilized to remove water for the methanolysis. Briefly, the samples were heated in 1 M methanolic HCl in a sealed screw-top glass test tube for 18 h at 80 °C. After cooling and removal of the solvent under a stream of nitrogen, the sample was treated with a 2:1:1 ratio mixture of methanol, pyridine, and acetic anhydride at room temperature for 30 min. The solvents were evaporated, and the sample was derivatized with 200 µL Tri-Sil (Pierce) at 80 °C for 30 min. Following extraction with hexane, GC/MS analysis of TMS methyl glycosides was performed on an Agilent 7890A GC interfaced to a 5975C MSD (mass selective detector) using a Supelco Equity-1–fused silica capillary column (30 m × 0.25 mm ID).
4
Glycosyl Composition Analysis by GC/MS
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Glycosyl composition analysis was performed by combined gas chromatography/mass spectrometry (GC/MS) of the per-O-trimethylsilyl (TMS) derivatives of the monosaccharide methyl glycosides produced from the sample by acidic methanolysis as described previously [48 ]. Briefly, the samples were heated with methanolic HCl in a sealed screw-top glass test tube for 18 h at 80°C. After cooling and removal of the solvent under a stream of nitrogen, the samples were treated with a mixture of methanol, pyridine, and acetic anhydride for 30 min. The solvents were evaporated, and the samples were derivatized with Sylon HTP (Sigma) at 80°C for 30 min. GC/MS analysis of the TMS methyl glycosides was performed on an Agilent 7890A GC interfaced to a 5975C MSD, using an Supelco Equity-1 fused silica capillary column (30 m × 0.25 mm ID).
5
Monosaccharide Composition Analysis by GC-MS
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Monosaccharide composition analysis was performed by combined GC-MS of the per-O-trimethylsilyl (TMS) derivatives of the monosaccharide methyl glycosides produced from the sample by acidic methanolysis as described previously73 (link). Briefly, released N-linked glycans were heated with methanolic HCl in a sealed screw-top glass test tube for 18 h at 80 °C. After cooling and removal of the solvent under a stream of nitrogen, the samples were re-N-acetylated at room temperature using methanol: pyridine: acetic anhydride (2:1:1) and dried again. The sample was then derivatized with Tri-Sil® (Pierce) at 80 °C for 20 min. GC/MS analysis of the TMS methyl glycosides was performed on an Agilent 7890A GC interfaced to a 5975 C MSD, using an Supelco Equity-1 fused silica capillary column (30 m × 0.25 mm ID). Inositol was added to the samples as an internal standard prior to data collection.
6
Glycosyl Composition Analysis by GC/MS
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Glycosyl composition analysis was done using gas chromatography/mass spectrometry (GC/MS) of the per-O-trimethylsilyl (TMS) derivatives of the monosaccharide methyl glycosides produced from the sample by acidic methanolysis (45) . The chloroform-treated whole peel (110 µg) was heated with methanolic HCl for 17 h at 80°C in a screw-top glass test tube. The sample had particulates floating in the solution after methylation, potentially cuticular material. The samples were cooled and the liquid was dried off using a stream of nitrogen. The tube was then treated by adding a mixture of methanol, pyridine, and acetic acid anhydride for 30 minutes.
Solvent was evaporated and the sample was derivatized using Tri-Sil (Pierce, USA) at 80°C for 30 minutes. TMS methyl glycosides were then analyzed on an Agilent 7890A GC interfaced to a 5975C MSD using an Equity-1 fused silica capillary column (30 m 0.25 mm ID, Supelco, USA).
Solvent was evaporated and the sample was derivatized using Tri-Sil (Pierce, USA) at 80°C for 30 minutes. TMS methyl glycosides were then analyzed on an Agilent 7890A GC interfaced to a 5975C MSD using an Equity-1 fused silica capillary column (30 m 0.25 mm ID, Supelco, USA).
7
Glycosyl Composition Analysis by GC/MS
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Glycosyl composition analysis was done using gas chromatography/mass spectrometry (GC/MS) of the per-O-trimethylsilyl (TMS) derivatives of the monosaccharide methyl glycosides produced from the sample by acidic methanolysis [45 (link)]. The chloroform-treated whole peel (110 µg) was heated with methanolic HCl for 17 h at 80 °C in a screw-top glass test tube. The sample had particulates floating in the solution after methylation, potentially cuticular material. The samples were cooled and the liquid was dried off using a stream of nitrogen. The tube was then treated by adding a mixture of methanol, pyridine, and acetic acid anhydride for 30 min. Solvent was evaporated and the sample was derivatized using Tri-Sil (Pierce, USA) at 80 °C for 30 min. TMS methyl glycosides were then analyzed on an Agilent 7890A GC interfaced to a 5975C MSD using an Equity-1 fused silica capillary column (30 m 0.25 mm ID, Supelco, USA).
8
Carbohydrate Composition and Linkage Analysis
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GC-MS analysis of the TMS methyl glycosides
and partially methylated alditol acetates (PMAAs) was performed using
an Agilent 7890A GC interfaced to a 5975C MSD. The composition analysis
employed a Supelco Equity-1 fused silica capillary column (30 m ×
0.25 mm ID), while the linkage analysis used a Supelco SP-2330 fused
silica capillary column (30 m × 0.25 mm ID). More details are
found in the Supporting Information.
and partially methylated alditol acetates (PMAAs) was performed using
an Agilent 7890A GC interfaced to a 5975C MSD. The composition analysis
employed a Supelco Equity-1 fused silica capillary column (30 m ×
0.25 mm ID), while the linkage analysis used a Supelco SP-2330 fused
silica capillary column (30 m × 0.25 mm ID). More details are
found in the Supporting Information.
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