HPLC/MS/MS with In-Source Collision-Induced Dissociation for Direct Measurement of PEGylated Compounds in Biological Matrices

摘要

Extraction and quantification of PEGylated compounds from biological matrices represents a major challenge. We report the validation of a bioanalytical method for the determination of small molecules conjugated to polyethylene glycols (PEG's) in plasma by liquid chromatography-atmospheric pressure ionization/tandem mass spectrometry (LC-API/MS/MS) detection. A PEGylated small molecule analog was used as the internal standard. The lower limit of quantitation (LLOQ) for the analyte was 10.0 ng/Ml. The calibration curves were acceptable lover a range of 10.0 - 1000 ng/ML. The analyte and internal standard were extracted from 100uL of plasma by protein precipitation by addition of acetonitrile containing 0.1percent formic acid (FA). After evaporation to dryness and reconstitution, the extracts were analyzed by LC-API/MS/MS. Run times were approximately 4.5 minutes. Because of the nature of this compound (PEGylated), relaxed acceptance criteria were set for the validation (+-20percent).