Angiotensin peptides (ANGs) play a central role in the renin–angiotensin–aldosterone system, rendering them interesting biomarkers associated with hypertension. Precise quantification of circulating ANGs holds the potential to assess the activity of angiotensin-converting enzyme (ACE), a key protease targeted by widely prescribed drugs, namely ACE inhibitors. This ability could pave the way for personalised medicine, offering insights into the prescription of inhibitors targeting either the proteases or the receptors within the system. Despite recent developments in liquid chromatography–mass spectrometry (LC-MS) methods for measuring circulating ANG concentrations, comprehensive stability studies of ANGs in human plasma are absent in the literature, raising concerns about the reliability of measured concentrations and their link to clinical conditions. To address this critical gap, we conducted an exhaustive evaluation of the pre-analytical stability of ANG1–10, ANG1–9, ANG1–8, ANG1–7, and ANG1–5. By employing surfactants to mitigate non-specific adsorption and a dedicated mix of protease inhibitors to limit protease activity, we established an MS-based assay for these five peptides. We used this method to quantify circulating concentrations of ANGs in the plasma of 11 healthy donors and 3 patients under kidney dialysis. Our findings revealed that ANG1–10 and ANG1–8 circulate at concentrations ranging from 1 to 10 pM in healthy subjects and exhibit a high degree of correlation. Notably, ANG1–9, ANG1–7, and ANG1–5 were undetectable in any of the 14 patients, despite a sub-picomolar limit of detection. This strikingly contrasts with the reference concentrations reported in the literature, which typically fall within the picomolar range. In light of these discrepancies, we strongly advocate for rigorous pre-analytical considerations and comprehensive stability studies to ensure reliable results. We emphasise the pivotal role of heightened pre-analytical awareness within the clinical chemistry community, and we hope for continued growth in this critical area.

中文翻译：

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使用微型 UHPLC-MS/MS 定量人血浆中的内源性血管紧张素 1-10、1-9、1-8、1-7 和 1-5：概述预分析对于获得可靠结果的重要性

血管紧张素肽（ANG）在肾素-血管紧张素-醛固酮系统中发挥着核心作用，使其成为与高血压相关的有趣的生物标志物。循环 ANG 的精确定量有可能评估血管紧张素转换酶 (ACE) 的活性，ACE 是广泛处方药物（即 ACE 抑制剂）所针对的关键蛋白酶。这种能力可以为个性化医疗铺平道路，为针对系统内蛋白酶或受体的抑制剂处方提供见解。尽管最近在测量循环 ANG 浓度的液相色谱-质谱 (LC-MS) 方法方面取得了进展，但文献中缺乏对人血浆中 ANG 的全面稳定性研究，这引起了人们对测量浓度的可靠性及其与临床状况的联系的担忧。为了解决这一关键差距，我们对 ANG1-10、ANG1-9、ANG1-8、ANG1-7 和 ANG1-5 的分析前稳定性进行了详尽的评估。通过使用表面活性剂来减轻非特异性吸附，并使用蛋白酶抑制剂的专用混合物来限制蛋白酶活性，我们建立了针对这五种肽的基于质谱的测定方法。我们使用这种方法量化了 11 名健康捐献者和 3 名肾透析患者血浆中 ANG 的循环浓度。我们的研究结果表明，健康受试者中 ANG1-10 和 ANG1-8 的循环浓度范围为 1 至 10 pM，并表现出高度相关性。值得注意的是，尽管检测限低于皮摩尔水平，但在 14 名患者中均检测不到 ANG1-9、ANG1-7 和 ANG1-5。这与文献中报道的参考浓度形成鲜明对比，文献中的参考浓度通常落在皮摩尔范围内。鉴于这些差异，我们强烈主张进行严格的分析前考虑和全面的稳定性研究，以确保结果可靠。我们强调临床化学界提高分析前意识的关键作用，我们希望这一关键领域能够持续增长。