Glu1 fibrinopeptide b

The fractions that showed antimicrobial activity were analyzed with a Nano Acquity UPLC system coupled to a quadropole time-of-flight mass spectrometer (Qtof Micro, Waters, Milford, MA). During MS analysis the capillary voltage was 4100 V, sample cone 30 V, and the collision energy was set to 5 V, but increased to 15, 25, 40, 55, and 65 V with a 1 second interscan time during MS/MS analysis. The column was a C18 Nano Acquity 75 μm × 250 mm column with 1.7 μm particle size (Waters, Milford, MA). The lock mass was a 1.27 μM solution of (Glu¹)-Fibrinopeptide B (Sigma Aldrich, St Louis, MO), infused at a flow rate of 0.4 μL/min. All spectra and chromatograms were analyzed with MassLynx version 4.0 software package. The two eluents: A and B, were water and acetonitrile, respectively, with 0.1% formic acid at a flow rate of 0.250 μL/min. All samples were injected and separated on the column with an isocratic step of 1% B for 1 minute, and a gradient increasing to 90% B during 49 minutes, and held for 4 minutes. To avoid possible microbial growth in eluent A, 0.05% MeOH was added.
¹H NMR experiments were performed on a Bruker 600 MHz spectrometer equipped with a smartprobe, using tetramethylsilane as internal standard. All samples were dissolved in 99.9% dimethylsulfoxide-d6 and analyzed at 25°C.

A comprehensive range of chemicals, including PHE, dithiothreitol (DTT), formic acid, [Glu1]-fibrinopeptide B, microbiological agar, and others essential for our procedures, were sourced primarily from Sigma-Aldrich (St. Louis, MO, USA) and Fluka (Buchs, Switzerland). Additional reagents, such as the complete mini-protease inhibitor cocktail, were procured from Roche (Mannheim, Germany), while LiChrosolv quality solvents were obtained from Merck (Darmstadt, Germany). Specific enzymes, including trypsin and benzonase, were supplied by Promega (Madison, WI, USA).