Kinetex 2.6 m c18 100a column

For reaction progress testing, liquid chromatography (HPLC) was performed using a KNAUER apparatus equipped with a UV-Vis detector with application of the standard procedure [70 (link),71 (link),72 (link)]. LiChrospher RP-18 10 μm column (4 × 250 mm) was applied and methanol–water MeOH:H₂O (70:30 v/v) was used as eluent at a flow rate of 1.5 cm³ min⁻¹. Melting points were determined with the Boetius PHMK 05 apparatus and were not corrected. IR spectra (KBr pellets) were registered on a Thermo Nicolet 6700 FT-IR apparatus. ¹H NMR (500 MHz) and ¹³C NMR (125 MHz) spectra were recorded with a Bruker AVANCE NMR spectrometer using CDCl₃ as a solvent. TMS was used as an internal standard. UV-Vis spectra were determined for the 200–500 nm range through usage of spectrometer UV-5100 Biosens. HR-MS spectra were performed on a Shimadzu LCMS-IT-TOF instrument with ES ionization (heat bock and CDL temperatures of 200 °C, nebulizing gas flow of 1.5 mL/min) connected to a Shimadzu Prominence two LC-20AD pump chromatograph equipped with Phenomenex Kinetex 2.6 µm C18 100A column (acetonitrile–water mixtures, ACN:H₂O, 65:35 v/v, were used as the eluent).

Spinal white matter was collected from three affected cats from the 1997 outbreak and six affected cats from the outbreak in 2001 as well as from the three control cats. Spinal cord material was frozen in liquid nitrogen and stored at −70°C until use. Immediately prior to analysis the material was thawed and ground. Also, a sample of the irradiated food associated with the 2000 outbreak (SSNIFF, 2.5Mrad = 25 kGy) and of a standard dry cat food from the same manufacturer were obtained and ground prior to analysis.
Cholesteryl esters (CE) were isolated by a modified “Bligh & Dyer” extraction according to Retra et al. (2008), followed by a solid‐phase extraction method according to Hamilton and Comai (1988). Thereafter the CEs were saponified according to the modified method described by Kates (1986), where petroleum ether was replaced by hexane. Polyunsaturated fatty acid analysis was performed by high‐pressure liquid chromatography/mass spectrometry (HPLC/MS) according to Retra et al. (2008) in which the Synergi 4 µm MAX‐RP 18A column was replaced by a Kinetex 2.6 µm C18 100A column (150 × 3 mm; Phenomenex, CA, USA). Internal standards were used for comparison.