Laboratory of Molecular & Evolutionary Parasitology, RAPID Group, School of Biosciences, University of Kent, Canterbury CT2 7NJ, UK.
Wellcome Trust Biomolecular NMR Facility, School of Biosciences, University of Kent, Canterbury CT2 7NJ, UK.
Molecules. 2021 May 29;26(11):3285. doi: 10.3390/molecules26113285.
is an opportunistic parasite commonly found in the intestines of humans and other animals. Despite its high prevalence, knowledge regarding biology within and outside the host is limited. Analysis of the metabolites produced by this anaerobe could provide insights that can help map its metabolism and determine its role in both health and disease. Due to its controversial pathogenicity, these metabolites could define its deterministic role in microbiome's "health" and/or subsequently resolve ' potential impact in gastrointestinal health. A common method for elucidating the presence of these metabolites is through H nuclear magnetic resonance (NMR). However, there are currently no described benchmarked methods available to extract metabolites from for H NMR analysis. Herein, several extraction solvents, lysis methods and incubation temperatures were compared for their usefulness as an extraction protocol for this protozoan. Following extraction, the samples were freeze-dried, re-solubilized and analysed with H NMR. The results demonstrate that carrying out the procedure at room temperature using methanol as an extraction solvent and bead bashing as a lysis technique provides a consistent, reproducible and efficient method to extract metabolites from for NMR.
是一种机会主义寄生虫，通常存在于人类和其他动物的肠道中。尽管它的流行率很高，但人们对宿主内外的生物学知识知之甚少。分析这种厌氧菌产生的代谢物可以提供深入的了解，有助于绘制其代谢图谱，并确定其在健康和疾病中的作用。由于其有争议的致病性，这些代谢物可以定义其在微生物组“健康”中的决定性作用，以及随后解决胃肠道健康方面的“潜在影响”。阐明这些代谢物存在的常用方法是通过 H 核磁共振（NMR）。然而，目前尚无描述的基准方法可用于从 中提取代谢物进行 H NMR 分析。本文比较了几种提取溶剂、裂解方法和孵育温度，以评估它们作为提取这种原生动物代谢物的方案的有用性。提取后，将样品冻干、复溶并进行 H NMR 分析。结果表明，在室温下使用甲醇作为提取溶剂，采用珠磨法作为裂解技术，可以提供一种从 中提取代谢物用于 NMR 的一致、可重复和有效的方法。
