Ng Josephine Su Yin, Ryan Una, Trengove Robert D, Maker Garth L
School of Veterinary and Biomedical Science, Murdoch University, Murdoch, Western Australia 6150, Australia.
Mol Biochem Parasitol. 2012 Oct;185(2):145-50. doi: 10.1016/j.molbiopara.2012.08.006. Epub 2012 Sep 1.
Faecal metabolite profiling, though in its infancy, allows for investigation of complex metabolic interactions between gastrointestinal infections or diseases and host health. In the present study, we describe a faecal metabolite extraction method for untargeted gas chromatography-mass spectrometry (GC-MS) analysis using Cryptosporidium positive and negative human faecal samples. The extraction method takes into account the varying faecal consistencies and quantities received for clinical diagnosis. Optimisation was carried out using different extraction solvents and on three different faecal quantities to determine the minimum amount of faecal sample required. The method was validated by untargeted GC-MS analysis on 8 Cryptosporidium positive and 8 Cryptosporidium negative human faecal samples, extracted using the optimised conditions. The method showed good extraction reproducibility with a relative standard deviation of 9.14%. Multivariate analysis of the GC-MS generated dataset showed distinct differences between profiles of Cryptosporidium positive and Cryptosporidium negative samples. The most notable differences included changes in amino acid, nitrogen and energy metabolism, demonstrating the association of infection with Cryptosporidium and altered permeability of the small intestine.
粪便代谢物谱分析尽管尚处于起步阶段，但可用于研究胃肠道感染或疾病与宿主健康之间复杂的代谢相互作用。在本研究中，我们描述了一种使用隐孢子虫阳性和阴性人类粪便样本进行非靶向气相色谱 - 质谱（GC-MS）分析的粪便代谢物提取方法。该提取方法考虑了临床诊断中收到的粪便稠度和数量的变化。使用不同的提取溶剂并针对三种不同的粪便量进行了优化，以确定所需的最小粪便样本量。该方法通过对8份隐孢子虫阳性和8份隐孢子虫阴性人类粪便样本进行非靶向GC-MS分析进行验证，并采用优化条件进行提取。该方法显示出良好的提取重现性，相对标准偏差为9.14%。对GC-MS生成数据集的多变量分析显示，隐孢子虫阳性和阴性样本的谱图存在明显差异。最显著的差异包括氨基酸、氮和能量代谢的变化，表明感染隐孢子虫与小肠通透性改变之间的关联。
