Mature biofilms were incubated in fresh media. After 16 hours, spent media was collected and diluted 1:1 in pure methanol. Methanolic extracts were separated by Ultra High-Performance Liquid Chromatography (UHPLC) mass spectrometry (MS), using a hydrophilic interaction liquid chromatography column (Syncronis HILIC, Thermo Fisher, stationary phase).52 (link) Solvent A (20mM ammonium formate pH 3 in H20) and B (MS Grade Acetonitrile with 0.1% formic acid) were used for LC Gradient. High-resolution full-scan MS data were acquired on a Thermo Fisher Scientific Q-Exactive HF mass spectrometer using negative-mode electrospray ionization. Metabolites were identified using retention times of standards using MAVEN freeware.53 (link)
Syncronis hilic
Manufactured by Thermo Fisher Scientific
The Syncronis HILIC is a high-performance liquid chromatography (HPLC) column designed for the separation of polar and hydrophilic compounds. It utilizes the hydrophilic interaction liquid chromatography (HILIC) principle to enable the analysis of a wide range of analytes, including amino acids, carbohydrates, and other water-soluble species.
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2 protocols using syncronis hilic
1
Metabolite Profiling of Mature Biofilms
2
Optimizing Bile Acids Separation
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Several columns including a Thermo Scientific Accucore™ 150 Amide HILIC (100 × 2.1 mm; 2.6 µm) column, a Thermo Scientific Syncronis™ HILIC (100 × 2.1 mm; 1.7 µm) column, an ACE C18 PFP (100 × 2.1 mm; 1.7 µm) column, an ACE C18 AR (100 × 2.1 mm; 1.7 µm) column, a Thermo Scientific Hypersil GOLD™ aQ (100 × 2.1 mm; 1.9 µm) column and an ACE C18 (100 × 2.1 mm; 1.7 µm) column were tested to obtain the best separation of the targeted bile acids. All reversed phase stationary phases were tested with water and ACN containing 0.1% formic acid as the mobile phase. HILIC stationary phases were tested with ACN: water containing 15 mM ammonium acetate adjusted to pH 5 with acetic acid as a mobile phase.
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