Approach
To achieve the LLOQ of 10 pg/mL all three components of the method; analyte isolation, separation and detection were optimised to be highly selective and sensitive.
Analyte Isolation: Due to the instability of the metabolites all plasma samples were stabilised in acidic pH and a SPE extraction developed using µ-elution SPE format which allowed the sample to be processed without the use of sample concentration using temperature or pressure.
Separation: Chromatographic separation was achieved by the use of sub 3 µm fused core column with a slow gradient system.
Detection: A Waters Xevo TQ-S LC-MS/MS system equipped with an electrospray interface was used for detection to reach the LLOQ and was approximately five times more sensitive than our AB SCIEX API 5000 instruments.