Luna rp c5 column

PEth was analyzed in 200 μL of whole blood by online-SPE-LC-MS/MS with a QTrap 3200 mass spectrometer (AB Sciex, Toronto, Canada). Analytical separation was performed by a Luna RP-C5 column, 50 mm x 2 mm, 5 μm particle size (Phenomenex, Brechbühler, Schlieren, Switzerland) by gradient elution using ammonium acetate (10mM)/MeCN (30:70, v/v) and 2-propanol. The precise details of the validated method can be found in [25] .

The online-SPE-LC-MS/MS system was composed of a CTC PAL autosampler (CTC Analytics, Zwingen, Switzerland), an Agilent 1200 series HPLC (Agilent, Waldbronn, Germany), a Hewlett Packard 1100 HPLC (Agilent, Waldbronn, Germany), and a QTrap 3200 mass spectrometer (Sciex, Toronto, Canada) controlled by Analyst 1.5.1 software.
Chromatographic separation was conducted with a Luna RP-C5 column, 50 mm × 2 mm, 5 μm (Phenomenex, Brechbühler, Schlieren, Switzerland) heated to 50 °C with a flow rate of 0.25 mL/min. The trapping column was a Synergi Polar-RP, 20 × 2 mm, 5 μm (Phenomenex, Brechbühler, Schlieren, Switzerland). Mobile phase A consisted of ammonium acetate (10 mM)/acetonitrile (30:70, v/v) and mobile phase B was 2-propanol. The mobile phase A for the trapping column consisted of 0.1 % HCOOH and acetonitrile (70:30, v/v); here, mobile phase B was also 2-propanol. PEth 16:0/ 18:1 and PEth 16:0/18:2 were separated with the following 12 min gradient: 0 to 2 min, 10 % B; 2 to 3.5 min, 10 to 99 % B linear; 3.5 to 6 min, 99 % B; 6 to 7.5 min, 99 to 10 % B linear; and 7.5 to 12 min, 10 % B with a retention time of 6.42 min for PEth 16:0/18:1 and 6.29 min for PEth 16:0/18:2.
The mass spectrometer was operated in ESI negative MRM mode, with an ion spray voltage of -4250 V and a source temperature of 650 °C with the following transitions:

The LC-MS/MS system consisted of a CTC PAL autosampler (CTC Analytics, Zwingen, Switzerland), an Agilent 1200 series HPLC (Agilent, Waldbronn, Germany) and a 3200 QTrap mass spectrometer (Sciex, Toronto, Canada) controlled by Analyst™ software (version 1.5.1).
Analytical separation was performed by a Luna RP-C5 column, 50 mm×2 mm, 5 μm (Phenomenex, Brechbühler, Schlieren, Switzerland) heated to 50 °C with a flow rate of 0.3 mL/min. Mobile phase A consisted of ammonium acetate (2 mM)/acetonitrile (30:70, v/v) solution, and mobile phase B was 2-propanol. The following 10-min gradient was used: 0 to 1.5 min, 10 % B; 1.5 to 2.5 min, 10 to 40 % B linear; 2.5 to 3.5 min, 40 to 100 % B linear; 3.5 to 4.5 min, 100 % B; 4.5 to 6 min, 100 to 10 % B linear; and 6 to 10 min, 10 % B. Postcolumn infusion of 2-propanol (0.3 mL/min) was used to increase the signal intensity.
The mass spectrometer was operated in negative ESI MRM mode, with an ion-spray voltage of -4250 V and a source temperature of 650 °C with the following transitions for PEth 16:0/18:1: m/z 701.5/255.1 (quantifier), m/z 701.5/ 281.1 and m/z 701.5/437.2 (qualifiers), and m/z 706.5/281.1 (D 5 -PEth 16:0/18:1). For PEth 16:0/18:2, the transitions were the following: m/z 699.5/255.2 (quantifier), m/z 699.5/279.2 and m/z 699.5/437.2 (qualifiers), and m/z 704.5/279.4 (D 5 -PEth 16:0/18:2).