The quality of sample extraction had a significant impact on mass spectrometry results. The presence of
phospholipids in the sample extracts resulted in poor quantitation and also it decreases the method robustness. Here
we adopted a novel sample preparation Hybrid SPE phospholipid technology to extract plasma samples for
improved phospholipid removal. This new method allowed simultaneous quantification of propafenone and 5-OH
propafenone at lower levels 0.5 and 0.25 ng/mL respectively. The phospholipid free filtrate obtained through Hybrid
SPE-Phospholipid cartridge was chromatographed onto Gemini C18 column (75 x 4.6 mm, 3.0 μm). An isocratic
mobile phase of a mixture of 10mM ammonium formate (pH 3.0 adjusted with formic acid) and methanol
(20:80%V/V) at a flow rate of 0.5 mL/min was used. Precursor ion and product ion transition for analytes and IS
were monitored on a TSQ Vantage triple quadrupole mass spectrometer, operated in the positive ionization mode.
Method was validated over a concentration range of 0.50-500.00 ng/mL for propafenone, 0.25-250.00 ng/mL for 5-
OH propafenone. The intra- and inter-day precision over the concentration range for propafenone and 5-OH
propafenone were lower than 6.1 and 14.2% (coefficient of variation, %CV), and accuracy was between 99.5–108.7
and 94.6–108.3%, respectively. By using this new Hybrid SPE-Phospholipid technology the risk of phospholipid
accumulation on column was knocked out completely and resulted in good peak shape with excellent column
performance.
