Exploring the role of ex vivo metabolism on blood and plasma measurements of oxytocin among women in the third stage of labour: A post hoc study

Aims: To examine the role of ex vivo oxytocin metabolism in post-dose peptide measurements. Methods: The stability of oxytocin (Study 1) and oxytocinase activity (Study 2) in late-stage pregnancy blood was quantified using liquid-chromatography tandem mass-spectrometry (LC–MS/MS) and a fluorogenic assay, respectively. Analyses were conducted using blood from pregnant women (>36 weeks gestation) evaluated in lithium heparin (LH), ethylenediaminetetraacetic acid (EDTA) and BD P100 blood collection tubes with or without protease inhibitors. In addition, plasma oxytocin concentrations following administration of oxytocin 240 IU inhaled, 5 IU intravenous or 10 IU intramuscular in women in third stage of labour (TSL) were analysed using enzyme-linked immunosorbent assay (ELISA) and LC–MS/MS to understand how quantified peptide concentrations differ between these analytical methods (Study 3). Results: Study 1: Oxytocin was stable in blood collected into EDTA tubes with or without protease inhibitors but not in LH tubes. Study 2: Blood collected into all EDTA-containing collection tubes led to near-complete inhibition of oxytocinase (≤100 min). In plasma, a 35% reduction in oxytocinase activity was observed in LH tubes with EDTA added. In plasma from late-stage pregnancy compared to nonpregnant participants, the oxytocinase activity was approximately 11-fold higher. Study 3: Plasma oxytocin concentrations from nonpregnant or women in TSL following exogenous oxytocin administration were ≤33 times higher when analysed using ELISA vs. LC–MS/MS methods. Conclusions: Collection of blood from late-stage pregnant women into tubes containing EDTA inhibits oxytocinase effectively stabilizing oxytocin, suggesting low concentrations of oxytocin after dose administration reflect rapid in vivo metabolism.