Department of Chemical and Geological Sciences, University of Cagliari, I-09042, Italy.
Department of Surgical Sciences, University of Cagliari, Italy.
Clin Chim Acta. 2018 Oct;485:236-242. doi: 10.1016/j.cca.2018.07.006. Epub 2018 Jul 6.
Before derivatization, urine analyzed by gas chromatography-mass spectrometry (GC-MS) requires the complete removal of urea to avoid interferences. We aimed at establishing the most effective sample pretreatment for urea removing; moreover, we explored the impact of two short-term sample storage conditions on urine metabolome.
92 aliquots were obtained from a single sample collected from a healthy adult; they were divided into 6 groups. Group 1 consisted of untreated aliquots while groups 2-6 differed from each other for the addition of various defined urease solution volumes combined with either 30 min or 1-hour sonication time. Urine sample storage was tested by comparing 20 fresh aliquots analyzed after collection with 20 aliquots frozen at -80 °C for 72 h.
the most effective protocol consisted of the combination between 200 μL urease solution with 1-h sonication time; urease solution volumes >200 μL increase the risk to underestimate metabolite peaks because of sample dilution. Short-term storage of samples at -80 °C pointed out significant changes in the urine metabolic profile compared with that of fresh samples.
our study confirms the importance of urea removal for a reliable recognition and quantitation of metabolites; urine short-term storage at -80 °C should be carefully reconsidered.
在衍生化之前，气相色谱-质谱联用（GC-MS）分析的尿液需要完全去除尿素以避免干扰。我们旨在建立最有效的尿素去除样品预处理方法；此外，我们还探讨了两种短期样品储存条件对尿液代谢组的影响。
从一名健康成年人的单个样本中获得 92 个等分试样；将它们分为 6 组。第 1 组为未处理的等分试样，而第 2-6 组则因添加不同体积的定义好的脲酶溶液以及 30 分钟或 1 小时的超声时间而有所不同。通过比较收集后立即分析的 20 个新鲜等分试样与在-80°C 下冷冻 72 小时的 20 个等分试样，测试了尿液样品的储存情况。
最有效的方案是 200μL 脲酶溶液与 1 小时超声时间的组合；脲酶溶液体积>200μL 会增加因样品稀释而低估代谢物峰的风险。与新鲜样品相比，-80°C 下的短期储存样品在尿液代谢谱方面发生了显著变化。
我们的研究证实了去除尿素对于可靠识别和定量代谢物的重要性；-80°C 下的尿液短期储存应谨慎重新考虑。
