N acetyl l tryptophan

Human serum albumin (HSA), essentially fatty acid free (A1887), phenylbutazone, taurine, N-acetyl-l-tryptophan, N-acetyl-l-tyrosine, methionine, cysteine, phenylalanine, alanine, serine, arginine, leucine, 5,5′-dithiobis-(2-nitrobenzoic acid) (DTNB), and 2,4-dinitrophenylhydrazine were purchased from Sigma-Aldrich (St. Louis, MO, USA). HSA was dissolved in 10 mmol·L⁻¹ phosphate-buffered saline (PBS) at pH 7.4 to give a 1 mmol·L⁻¹ stock solution, and its concentration was measured by its absorbance (ε_{280 nm} = 35,219 mol⁻¹·L·cm⁻¹). Hypochlorous acid (HOCl) was prepared by diluting a 5% stock solution, and the concentration was determined spectrophotometrically after dilution in 0.01 mol·L⁻¹ NaOH, pH 12 (ε_{292 nm} = 350 mol⁻¹·L·cm⁻¹). taurine monochloramine (Tau-NHCl) was prepared by the addition of 5 mmol·L⁻¹ HOCl to 50 mmol·L⁻¹ taurine in PBS at pH 7.4 [53 (link)]. Hypobromous acid (HOBr) was synthesized by combining 100 mmol·L⁻¹ HOCl and 200 mmol·L⁻¹ NaBr in water. taurine dibromamine (Tau-NBr₂) was prepared by the addition of 5 mmol·L⁻¹ HOBr to 2.5 mmol·L⁻¹ taurine in PBS at pH 7.4 [54 (link)]. A Perkin Elmer Lambda 35 UV–visible (UV–Vis) spectrophotometer (Shelton, CT, USA) was used for the UV–Vis measurements.

LC–MS grade methanol, water and formic acid as well as the standard mixtures used for the external calibration of the MS instrument were from Thermo Fisher Scientific (MA, USA). All analytical grade reference compound (glycocholic acid, d-tryptophan, N-acetyl-l-tryptophan, L-kynurenine and Kynurenic acid) were from Sigma Aldrich (Steinheim, Germany).

After injury, randomly selected animals in each injury group were administered with either 1 mg/kg of the lipid soluble NK1 antagonist EUC-001¹² (Eustralis Pharmaceuticals, Melbourne, Australia), 2.5 mg/kg of the non-lipid soluble NK1 antagonist n-acetyl L-tryptophan (Sigma-Aldrich, Castle Hill, Australia), or equal volume saline vehicle. Drugs were administered as a single bolus i.v. in rats and i.p. in mice at 30 min after single injury, or 30 min after each injury induction following repeated mild TBI.