Strategy for Nuclear-Magnetic-Resonance-Based Metabolomics of Human Feces

Lamichhane, Santosh; Yde, Christian Clement; Schmedes, Mette Søndergaard; Jensen, Henrik Max; Meier, Sebastián; Bertram, Hanne Christine

doi:10.1021/acs.analchem.5b00977

Cited by 68 publications

(60 citation statements)

References 33 publications

Supporting

Mentioning

Contrasting

Unclassified

Order By: Relevance

“…Very recently Lamichhane et al reported that the feces weight to extraction solvent volume (wf/vs) ratio has the strongest impact on the metabolic profile [41]. This parameter together with the buffer pH value were considered to be the most significant in our study in the process of fecal sample extraction prior to NMR and GC-MS and were evaluated on the basis of the acquired metabolite profiles.…”

Section: H Nmr Profiling Datamentioning

confidence: 97%

“…Till now only few studies have dealt with the optimisation of the method used for the sample preparation of feces. In these studies metabolomics analysis was performed by NMR spectroscopy [41][42][43] and GC-MS [44][45][46][47][48] and the investigated parameters included freeze-drying, sonication, filtration, homogenization, sample weight to buffer/solvent ratio, extraction solvent, pH value, extraction duration and repetitions numbers. Up to date there is no study of the optimum treatment conditions of fecal samples for metabolic fingerprinting by LC-MS.…”

mentioning

confidence: 99%

“…Wu et al[42] investigated, among other parameters, the extraction ratios of 1/7, 1/10/ 1/20 and 1/33 and suggested the 1/10 ratio as the optimum in neutral PBS for the NMR analysis of rat feces. Very recently Lamichhane et al,[41] examined human fecal samples extracted under 1/2, 1/5, 1/10 ratios and found that in the 1/2 wf/vs ratio the majority of metabolites were extracted more effectively. Our findings are in agreement with Lamichhane et al, however it should be noted that such conditions may be challenging for the practitioner.…”

mentioning

confidence: 99%

See 2 more Smart Citations

Sample preparation optimization in fecal metabolic profiling

Deda

Chatziioannou

Fasoula

et al. 2017

Journal of Chromatography B

View full text Add to dashboard Cite

Section: H Nmr Profiling Datamentioning

confidence: 97%

mentioning

confidence: 99%

mentioning

confidence: 99%

See 1 more Smart Citation

Sample preparation optimization in fecal metabolic profiling

Deda

Chatziioannou

Fasoula

et al. 2017

Journal of Chromatography B

View full text Add to dashboard Cite

“…Published studies so far have focused on developing various fecal extraction and sample preparation methods for metabolic profiling 12,13 . However, a standard protocol for fecal sample handling has yet to be established based on a systematic evaluation of experimental conditions.…”

mentioning

confidence: 99%

Optimized Sample Handling Strategy for Metabolic Profiling of Human Feces

Gratton¹,

Phetcharaburanin²,

Mullish³

et al. 2016

Anal. Chem.

143

127

View full text Add to dashboard Cite

Fecal metabolites are being increasingly studied to unravel the host-gut microbial metabolic interactions. However, there are currently no guidelines for fecal sample collection and storage based on a systematic evaluation of the effect of time, storage temperature, storage duration and sampling strategy. Here we derive an optimized protocol for fecal sample handling with the aim of maximizing metabolic stability and minimizing sample degradation. Samples obtained from five healthy individuals were analyzed to assess topographical homogeneity of feces, and to evaluate storage duration-, temperature-and freeze-thaw cycle-induced metabolic changes in crude stool and fecal water using a 1 H NMR spectroscopy-based metabolic profiling approach. Inter-individual variation was much greater than that attributable to storage conditions. Individual stool samples were found to be heterogeneous and spot sampling resulted in a high degree of metabolic variation. Crude fecal samples were remarkably unstable over time and exhibited distinct metabolic profiles at different storage temperatures. Microbial fermentation was the dominant driver in time-related changes observed in fecal samples stored at room temperature and this fermentative process was reduced when stored at 4°C. Crude fecal samples frozen at -20°C manifested elevated amino acids and nicotinate and depleted short chain fatty acids compared to crude fecal control samples. The relative concentrations of branched-chain and aromatic amino acids significantly increased in the freeze-thawed crude fecal samples, suggesting a release of microbial intracellular contents. The metabolic profiles of fecal water samples were more stable compared to crude samples. Our recommendation is that intact fecal samples should be collected, kept at 4°C or on ice during transportation, and extracted ideally within 1 h of collection, or a maximum of 24 h. Fecal water samples should be extracted from a representative amount (~15 g) of homogenized stool sample, aliquoted and stored at < -20°C, avoiding further freeze-thaw cycles.Metabolic profiling of biofluids and tissues generates data on a wide range of metabolites and provides extensive metabolic information on multiple biological processes in complex superorganisms such as mammals. Although urine and blood are often used to investigate systemic responses of animals and humans to various environmental stimuli or therapeutic interventions 1 , the search for disease biomarkers in fecal samples and studies on host-microbial interactions have intensified over the last decade. The human intestinal tract harbors >100 trillion microbial cells 2 and these microbes exert their influences on the human host primarily by metabolic signaling and therefore optimized methodologies for the study of microbial metabolic footprint is crucial to this field. Mounting evidence shows that the microbial composition and its collective metabolic activity profoundly impacts host physiology and modulates the disease risk of the host 3 . To investigate th...

show abstract

“…Fecal metabolites were extracted as described elsewhere [11] with some minor modifications. Briefly, the aliquots of about 100 mg thawed stool material were mixed with 0.5 mL phosphate buffer saline (1.9 mM Na 2 HPO 4 , 8.1 mM NaH 2 PO 4 , 150 mM NaCl, pH 7.4; Sigma-Aldrich, Germany) containing 10% deuterated water (D 2 O 99.8%; Cortecnet, France) and 0.05 mM sodium 3-trimethylsilyl-propionate- d 4 (TMSP-2,2,3,3- d 4 ; Cambridge Isotope Laboratories Inc., UK) as chemical shift reference.…”

Section: Methodsmentioning

confidence: 99%

1H-NMR analysis of feces: new possibilities in the helminthes infections research

et al. 2017

View full text Add to dashboard Cite

BackgroundAnalysis of the stool samples is an essential part of routine diagnostics of the helminthes infections. However, the standard methods such Kato and Kato-Katz utilize only a fraction of the information available. Here we present a method based on the nuclear magnetic resonance spectroscopy (NMR) which could be auxiliary to the standard procedures by evaluating the complex metabolic profiles (or phenotypes) of the samples.MethodThe samples were collected over the period of June-July 2015, frozen at −20 °C at the site of collection and transferred within four hours for the permanent storage at −80 °C. Fecal metabolites were extracted by mixing aliquots of about 100 mg thawed stool material with 0.5 mL phosphate buffer saline, followed by the homogenization and centrifugations steps. All NMR data were recorded using a Bruker 600 MHz AVANCE II spectrometer equipped with a 5 mm triple resonance inverse cryoprobe and a z-gradient system.ResultsHere we report an optimized method for NMR based metabolic profiling/phenotyping of the stools samples. Overall, 62 metabolites were annotated in the pool sample using the 2D NMR spectra and the Bruker Biorefcode database. The compounds cover a wide range of the metabolome including amino acids and their derivatives, short chain fatty acids (SCFAs), carboxylic acids and their derivatives, amines, carbohydrates, purines, alcohols and others. An exploratory analysis of the metabolic profiles reveals no strong trends associated with the infection status of the patients. However, using the penalized regression as a variable selection method we succeeded in finding a subset of eleven variables which enables to discriminate the patients on basis of their infections status.ConclusionsA simple method for metabolic profiling/phenotyping of the stools samples is reported and tested on a pilot opisthorchiasis cohort. To our knowledge this is the first report of a NMR-based feces analysis in the context of the helminthic infections.

show abstract

Strategy for Nuclear-Magnetic-Resonance-Based Metabolomics of Human Feces

Cited by 68 publications

References 33 publications

Sample preparation optimization in fecal metabolic profiling

Sample preparation optimization in fecal metabolic profiling

Optimized Sample Handling Strategy for Metabolic Profiling of Human Feces

1H-NMR analysis of feces: new possibilities in the helminthes infections research

Contact Info

Product

Resources

About