Department of Clinical Biochemistry, Glasgow Royal Infirmary, Glasgow, UK *Joint First author.
Ann Clin Biochem. 2021 Nov;58(6):622-631. doi: 10.1177/00045632211038021. Epub 2021 Aug 14.
Urinary 5-hydroxyindoleacetic acid (5-HIAA) is a first-line investigation for gastrointestinal neuroendocrine tumours that secrete serotonin. It also has clinical utility for monitoring disease progression and therapeutic response.
To develop and validate a liquid chromatography-tandem mass spectrometry (LC-MS/MS) method for urinary 5-hydroxyindoleacetic acid that incorporates a supported liquid extraction and C-labelled internal standard.
Samples were diluted in ammonium acetate containing a C-labelled internal standard (5-hydroxyindole-3a,4,5,6,7,7a-C-3-acetic acid). Supported liquid extraction was performed followed by chromatographic separation using the 2.1 × 30 mm CORTECS® UPLC® T3 column. Mass spectrometry detection (Waters Xevo TQ-XS) was performed in electrospray positive mode using the transitions 192.3 > 146.4 m/z (quantifier) and 192.3 > 118.4 m/z (qualifier) for 5-hydroxyindoleacetic acid and 198.2 > 152.4 m/z for C-5-HIAA.
A well-defined 5-hydroxyindoleacetic acid peak was observed at 0.8 min with a run time of 2.4 min. The assay was linear (r > 0.99) to 382 mol/L, with a lower limit of quantification of 5.3 mol/L (CV <15%). Analysis of 29 external quality assurance samples showed good agreement between our method and the UKNEQAS method mean (4.7% positive bias). The intra- and inter-assay precision was within acceptable limits, and the assay was stable up to 96 h postextraction with minimal carryover.
We have developed a robust LC-MS/MS method with semi-automated extraction that offers an improved run time and performance over the existing, labour-intensive, HPLC method. The method was quick, precise, showed good agreement with UKNEQAS external quality assurance material and is in routine service for clinical samples.
尿 5-羟吲哚乙酸（5-HIAA）是一种用于检测分泌血清素的胃肠道神经内分泌肿瘤的一线检查方法。它在监测疾病进展和治疗反应方面也具有临床应用价值。
开发和验证一种包含固相萃取和 C 标记内标物的液相色谱-串联质谱（LC-MS/MS）法测定尿 5-HIAA。
样品用含有 C 标记内标物（5-羟吲哚-3a、4、5、6、7、7a-C-3-乙酸）的乙酸铵稀释。采用固相萃取法，然后使用 2.1×30mm CORTECS®UPLC®T3 柱进行色谱分离。采用电喷雾正模式进行质谱检测（Waters Xevo TQ-XS），用于 5-HIAA 的定量和定性分析的转换分别为 192.3＞146.4m/z（定量器）和 192.3＞118.4m/z，用于 C-5-HIAA 的转换为 198.2＞152.4m/z。
在 0.8min 处观察到定义明确的 5-HIAA 峰，运行时间为 2.4min。该方法呈线性（r＞0.99）至 382μmol/L，定量下限为 5.3μmol/L（CV＜15%）。对 29 份外部质量保证样本的分析表明，本方法与 UKNEQAS 方法均值之间具有良好的一致性（正向偏差 4.7%）。内、日间精密度均在可接受范围内，萃取后 96h 内稳定，且无明显的交叉污染。
我们开发了一种稳健的 LC-MS/MS 方法，采用半自动提取，与现有的劳动强度大的 HPLC 方法相比，具有改进的运行时间和性能。该方法快速、精确，与 UKNEQAS 外部质量保证材料具有良好的一致性，并且已投入临床样本的常规使用。
