Poroshell hph c18 separation column

Amino acids were quantified in samples taken at half-maximal and maximal growth of 4 biological cultivations. Samples were prepared and analyzed by a 1260 Infinity HPLC system equipped with a fluorescence detector (Agilent Technologies, Waldbronn, Germany) and a Poroshell HPH-C18 separation column (4.6 x 100 mm, particle size 2.7 mm; Agilent Technologies) as described previously [21 ]. Samples for metabolome analysis were taken of 4 biological cultivations at half-maximal growth and each cultivation was analyzed in triplicates. Cell lysis, preparation and metabolome analysis was performed as described previously [18 (link), 22 (link)] with some minor modifications: ethanol was replaced by methanol and 500 μL of the polar phase were dried prior to derivatization.

High performance liquid chromatography measurements of free amino acids were performed on a 1260 Infinity HPLC system equipped with a fluorescence detector (Agilent Technologies, Waldbronn, Germany) and a Poroshell HPH-C18 separation column (4.6 mm × 100 mm, particle size 2.7 mm; Agilent Technologies). Before measuring, ammonium was precipitated from the samples by a 1:1 dilution with sodium tetraphenylborate (250 mM). After vigorously mixing and centrifugation (5 min, 13,000 rpm, 4°C), the ammonium-free samples were transferred in HPLC vials. The HPLC method was used as described previously (Trautwein et al., 2016 (link); Dannheim et al., 2017b (link)) with the minor modification that mobile phase A was changed to 25 mM sodium acetate (pH 6.5).