Abstract The quantitation of metanephrine (MN), normetanephrine (NMN), and 3-methoxytyramine (3MT) – referred to as metanephrines -- by LC-MS/MS is the gold-standard for screening for pheochromocytoma and paragangliomas (PPGLs), tumors of the adrenal gland and the peripheral nervous system. An assay for metanephrines from dried blood spots (DBSs) would be of high clinical utility as it simplifies sample collection, enables remote sampling, and could increase compliance with the clinical recommendation for supine sampling. Moreover, DBS sampling facilitates the measurement of blood-derived metanephrines in pediatric patients – where DBSs are well-established – in order to diagnose neuroblastomas. Here, we adapted an established derivatization-based LC-MRM-MS assay for plasma catecholamines, and optimized the sample extraction, LC, and MS parameters to produce a fast, sensitive, and robust method for the measurement of metanephrines from DBSs, including 3-methoxytyramine. The DBS samples were excised, derivatized with phenyl isothiocyanate (PITC) on-spot, extracted, and measured by LC-MRM-MS. To validate assay suitability and performance, we assessed the linearity, precision, accuracy, recovery, and matrix effects of the method, and determined the stability of metanephrines in DBSs under different storage conditions. Assay performance for NMN, MN, and 3MT was sufficient for quantitation from a single DBS within a linear range from 40 – 2000 pg/mL MN and NMN were stable in DBSs for 2 weeks, whereas 3MT was stable for one week regardless of storage temperature. Altogether, this work represents the first quantitative LC-MS/MS method for metanephrines from DBSs and provides a novel opportunity for the diagnosis of PPGLs and neuroblastomas in the future.

中文翻译：

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一种 LC-MRM 测定法，用于从干血斑中定量变肾上腺素，以诊断嗜铬细胞瘤和副神经节瘤

摘要 变肾上腺素 (MN)、去甲变肾上腺素 (NMN) 和 3-甲氧基酪胺 (3MT)（简称变肾上腺素）的定量 LC-MS/MS 是筛查嗜铬细胞瘤和副神经节瘤 (PPGL)、肿瘤的金标准肾上腺和周围神经系统。从干血斑 (DBS) 中检测变肾上腺素将具有很高的临床效用，因为它简化了样本收集，实现了远程采样，并且可以增加对仰卧采样的临床建议的依从性。此外，DBS 采样有助于在儿科患者中测量血源性变肾上腺素（DBS 已建立），以诊断神经母细胞瘤。在这里，我们采用了已建立的基于衍生化的 LC-MRM-MS 检测血浆儿茶酚胺，并优化了样品提取、LC、和 MS 参数，以产生一种快速、灵敏且可靠的方法，用于测量 DBS 中的变肾上腺素，包括 3-甲氧基酪胺。DBS 样品被切除，用异硫氰酸苯酯 (PITC) 现场衍生，提取，并通过 LC-MRM-MS 测量。为了验证测定的适用性和性能，我们评估了该方法的线性度、精密度、准确度、回收率和基质效应，并确定了不同储存条件下 DBS 中变肾上腺素的稳定性。NMN、MN 和 3MT 的测定性能足以在 40 – 2000 pg/mL 的线性范围内从单个 DBS 进行定量 MN 和 NMN 在 DBS 中稳定 2 周，而 3MT 稳定 1 周，无论储存温度如何. 共，