An optimised sample preparation method for NMR-based faecal metabonomic analysis
An optimised sample preparation method for NMR-based faecal metabonomic analysis
复制标题
基于 NMR 的粪便代谢组分析的优化样品制备方法
DOI：
10.1039/b927543f
复制
发表时间：
2010-01-01
期刊：
影响因子：
4.2
通讯作者：
Tang, Huiru
中科院分区：
文献类型：
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作者：
Wu, Junfang;An, Yanpeng;Tang, Huiru
粪便代谢组学核磁共振分析在研究哺乳动物代谢与共生肠道微生物群之间的相互作用中起着至关重要的作用。然而，粪便代谢物提取方法仍有待优化和标准化，以考虑信噪比、pH和化学位移一致性。在本研究中，我们比较了手动超声、组织酶自动均质及其组合三种均质方法的提取一致性，并系统优化了粪便代谢物提取参数，包括粪便与缓冲液比(W（f): V(b)）、提取次数和提取时间。我们发现，采用组织酶自动均质法提取代谢物具有良好的一致性和高通量。我们还建议W(f): V(b)为1:10 (mg μ l(-1))，并使用前两种提取物的组合作为合成样品来代表粪便代谢物的组成。这样的推荐是基于最大化光谱信噪比、pH值和化学位移一致性、代谢物提取的完整性和样品制备吞吐量的考虑，以便该方法适合分析大量样品，特别是在人群研究中。
Faecal metabonomic NMR analysis plays an essential role in investigating the interactions between mammalian metabolism and symbiotic gut microbiota. However, the faecal metabolite extraction method remains to be optimised and standardised to take into consideration signal-to-noise ratios, pH and chemical shift consistency. In the current investigation, we compared extraction consistency of three homogenisation methods including manual ultrasonication, automatic homogenization with tissuelyser and their combination, and systematically optimised faecal metabolite extraction parameters, including the faeces-to-buffer ratio (W(f) : V(b)), extraction repetition times and duration. We found that automatic homogenisation with tissuelyser was the choice of extraction method owning to its good metabolite extraction consistency and high throughput. We also recommend W(f) : V(b) of 1 : 10 (mg mu l(-1)) and use of the combined first two extracts as the resultant samples to represent faecal metabolite composition. Such recommendation is based on considerations of maximisation of the spectral signal-to-noise ratio, pH and chemical shift consistency, completeness of metabolite extraction and sample preparation throughput so that the method is suitable for analysing a large number of samples especially in human population studies.