Innovative Solutions - Small Molecule Bioanalysis

Unstable Metabolites

Quantification required for small molecule pro-drug, drug and two unstable metabolites to support clinical study. Target LLOQ: 10 pg/mL for all four analytes. Customer had been unsuccessful with developing the method at another bioanalytical CRO.


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.


Precise and accurate high sensitivity assay enabled the measurement of the complete PK profile of a high potency pro-drug, drug and its major metabolites.