DEVELOPMENT OF LC-MS/MS METHOD FOR DETERMINATION OF THE OLANZAPINE IN HUMAN PLASMA.

The study aim was to develop a validated selective, reproducible and sensitive Liquid Chromatography - Tandem Mass Spectrometry (LC-MS/MS) method for qualitative and quantitative determination of olanzapin (OLZ ) in human plasma. For isolation was used solid phase extraction with the Gilson GX-271 ASPEC Solid phase extraction system, cartridge Supelclean LC-18 SPE 500mg. Elution solvent contained 40mL of isopropanol, 4mL of concentrated ammonium hydroxide and 156mL of freshly prepared methylene chloride. Samples containing OLZ and IS were chromatographed on AGILENT TECHNOLOGIES 1290 Infinity AGILENT TECHNOLOGIES 6460 Triple quad LC-MS/MS system. Separation was performed by isocratic elution on Zorbax Eclipse plus C18 (100´4.5 mm, 3.8 µm) column, equipped with pre-column: UHPLC GUARD Zorbax Eclipse plus C18 (5×2.1 mm, 3.8 µm); column temperature was 35°C. The mobile phases consisted of 0.1 % water solution of formic acid HCOOH (H2O):0.1 % acetonitrile solution of formic acid HCOOH (CH3CN) in 70 :30 (v/v) proportion. The flow rate was 0.4 mL/min. Determination was performed on a triple-quadrupole mass spectrometer employing electrospray ionization technique (ESI+), operating in multiple reaction monitoring (MRM) and positive ion mode. Total chromatographic run time was 5 min. The optimized MRM transitions for OLZ were m/z 313 ® 256, m/z 313 →213 and m/z 313→198 as the quantitative and confirmative traces, respectively. Method was validated on following parameters: specificity, linearity, LLOQ, accuracy and precision, recovery, matrix effect and stability. The method was subsequently applied to in vitro plasma samples for determination of OLZ. The obtained data allows the opportunity for forensic investigation of the intoxication in postmortem cases.