Abstract:
OBJECTIVE: Postpartum haemorrhage (PPH) is the leading cause of maternal mortality worldwide. To prevent PPH, the WHO recommends administration of oxytocin (OT) immediately after birth, i.e. during the third stage of labour (TSL). Previous studies demonstrate that methods to quantify OT in biological matrices, e.g. enzyme-linked immunosorbent assays (ELISA), radioimmunoassays (RIA) and liquid chromatography-tandem mass spectrometry (LC-MS/MS) lack the specificity and/or sensitivity to accurately quantify OT in plasma from women administered OT during TSL. This is due to increased metabolic clearance of OT in late-stage pregnancy and at the time of childbirth, resulting in extremely low OT plasma concentrations. This study describes the development of an ultra-sensitive bioanalytical method that overcomes the issues previously reported and enables accurate pharmacokinetic analyses of exogenously administered OT in TSL.
METHODS: A selective and sensitive assay to quantify OT in TSL plasma was developed. Immunoprecipitation (IP) was applied to selectively extract OT from the TSL plasma, thereby generating clean extracts compatible with nanoflow LC (nLC). nLC-MS/MS was chosen for its high sensitivity and ability to differentiate between OT and potentially co-captured OT-like immunoreactive products.
RESULTS: The presented methodology is accurate and precise, with a good linear fit between 100-10 000 fg mL-1 OT. TSL plasma samples from a clinical phase 1 study (NCT02999100) were analysed successfully, enabling OT quantification down to 100 fg mL-1.
CONCLUSIONS: The presented IP-nLC-MS/MS method succeeded in overcoming the sensitivity challenge related to the assay of OT in TSL plasma and thereby revealing the PK profiles of OT in TSL plasma clinical study samples.
摘要:
目的:产后出血(PPH)是全球孕产妇死亡的主要原因。为了防止PPH,世卫组织建议在出生后立即给予催产素(OT),即在第三劳动阶段(TSL)。以前的研究表明,量化生物基质中OT的方法,例如酶联免疫吸附测定(ELISA),放射免疫分析(RIA)和液相色谱-串联质谱(LC-MS/MS)缺乏特异性和/或敏感性,无法准确定量在TSL期间服用OT的女性血浆中的OT。这是由于妊娠晚期和分娩时OT的代谢清除率增加,导致极低的OT血浆浓度。这项研究描述了一种超灵敏的生物分析方法的开发,该方法克服了先前报道的问题,并能够对TSL中外源施用的OT进行准确的药代动力学分析。
方法:开发了一种选择性和灵敏的方法来定量TSL血浆中的OT。免疫沉淀法(IP)用于从TSL血浆中选择性提取OT,从而产生与纳米低LC(nLC)相容的清洁提取物。选择nLC-MS/MS是因为其高灵敏度和区分OT和潜在共捕获的OT样免疫反应产物的能力。
结果:所提出的方法是准确和精确的,在100-10000fgmL-1OT之间具有良好的线性拟合。成功分析了临床1期研究(NCT02999100)的TSL血浆样本,使OT定量低至100fgmL-1。
结论:提出的IP-nLC-MS/MS方法成功地克服了与TSL血浆中OT测定相关的敏感性挑战,从而揭示了TSL血浆临床研究样品中OT的PK谱。