Development of a selective clean‐up method using immobilized antibody and its application to HPLC and GC/MS determination of a carbacyclin derivative, CS‐570, in plasma

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Abstract
Methods for the determination of CS‐570, a chemically stable prostacyclin analogue, in plasma were developed by using an immobilized antibody column followed by fluorescence HPLC and GC/MS. The CS‐570 antibody, obtained from rabbit plasma by giving CS‐570–BSA for a few months, was coupled to Sepharose 4B and used as extraction phase for sample clean‐up and extraction of the drug. A plasma sample was applied to this column, washed with water and the drug was eluted with 90% acetonitrile. 0.02% (w/v) 9‐anthryldiazomethane (ADAM) was added to the extract to form a fluorescent derivative. The CS‐570–ADAM adduct exhibited high sensitivity when applied to HPLC with fluorescence detection and column switching. The detection limit was 1 ng/mL when 1 mL of plasma was available. Additionally, a pentafluorobenzyltrimethylsilyl derivative of CS‐570 showed excellent sensitivity when determined by capillary GC interfaced to negative ion chemical ionization MS using a stable isotope labelled analogue as an internal standard.