Rapid and reliable HILIC-MS/MS method for monitoring allantoin as a biomarker of oxidative stress
Rapid and reliable HILIC-MS/MS method for monitoring allantoin as a biomarker of oxidative stress
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DOI：
10.1016/j.ab.2019.113509
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发表时间：
2020-01-15
影响因子：
2.9
通讯作者：
Kalikova, Kveta
中科院分区：
文献类型：
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作者：
Kozlik, Petr;Hasikova, Lenka;Kalikova, Kveta
尿囊素是人体氧化应激的一种极好的生物标志物，是活性氧氧化尿酸的主要产物。然而，尿囊素测定仍然没有在临床实验室中常规进行。因此，我们开发了一种快速，简单，选择性，灵敏度高的超高效液相色谱-串联质谱法测定人血清中的尿囊素使用同位素标记的内标物。我们的分析方案提供了高灵敏度的质谱检测和高通量的HILIC-MS/MS分析在4分钟内，一步血清样品制备约在7分钟内。最后，我们的协议进行了充分验证，证明其可靠性在人血清中的尿囊素测定。方法线性范围为0.05 ~ 100 μ M，精密度为1.8 ~ 11.3%（RSD），准确度（相对误差%）在± 6.0%以内。然后，该方法被应用于分析血清样品中的尿囊素浓度从71例慢性痛风没有治疗黄嘌呤氧化酶抑制剂。队列中的血清尿囊素浓度中位数为2.8 μ M（n = 71）。总的来说，我们简单的分析方案有可能很容易地在临床常规实践中实施，以监测尿囊素作为一个关键的氧化应激生物标志物。
Allantoin is an excellent biomarker of oxidative stress in humans as the main product of uric acid oxidation by reactive oxygen species. Yet, allantoin determination is still not routinely performed in clinical laboratories. Therefore, we developed a fast, simple, selective, and sensitive UHPLC-MS/MS method for allantoin determination in human serum using an isotopically labeled internal standard. Our analytical protocol provided high sensitivity by mass spectrometry detection and high throughput by HILIC-MS/MS analysis within 4 min, with one-step serum sample preparation approximately within 7 min. Lastly, our protocol was fully validated to demonstrate its reliability in allantoin determination in human serum. The method showed an excellent linear range from 0.05 to 100 mu M, with precision ranging from 1.8 to 11.3% (RSD), and with accuracy (relative error %) within +/- 6.0%. The method was then applied to analyze the concentration of allantoin in serum samples from 71 patients with chronic gout without treatment with xanthine oxidase inhibitors. The median serum allantoin concentration in the cohort was 2.8 mu M (n = 71). Overall, our simple analytical protocol has the potential to be easily implemented in clinical routine practice for monitoring allantoin as a key oxidative stress biomarker.