Utilizing NMR fecal metabolomics as a novel technique for detecting the physiological effects of food shortages, chronic stressors, in waterfowl
Description
NMR Spectra for Utilizing NMR fecal metabolomics as a novel technique for detecting the physiological effects of food shortages, chronic stressors, in waterfowl
Files
Steps to reproduce
All fecal samples were collected in cryovials and flash frozen in the field in liquid ni-trogen to stop metabolite degradation. Samples were taken to the lab to be stored at -80C until sample analysis. To remove water and standardize sample weights, fecal samples were lyophilized in a Labconco Freezone (Labcono Corporation, Kansas City, MO USA). 10 mg of freeze-dried fecal powder was extracted according to previously published protocols in chickens using 500 µl of 0.2M pH7.4 + NaN3 3mM +TSMP 0.1% phosphate buffer in D2O with 200 µl of supernatant pipetted into a 5mm NMR stem tubes. [31] Extracted samples were subjected to non-targeted 1H NMR spectroscopy at a Bruker Advance III HD 600 MHz NMR spectrometer. A standard 1- dimension-al noesypr1d pulse sequence (noesypr1d, 90˚ pulse length 15 µs, total acquisition time of 4 s) with water suppression applied during relaxation delay (1 s) and mixing time (100 ms) at 298 k. For each sample, 128 scans were recorded. RAW NMR spectra were processed Topspin 3.6 (Bruker Corporation, MA, USA) and then in Che-nomx NMR Suite 7.0 (Chenomx, Edmonton, Canada) for baseline and shimming cor-rection. Chenomx was then used to identify and measure concentrations of compounds visible in the NMR spectra. Concentrations of compounds present were used for statis-tical analysis in MetaboAnalyst 5.0 (https://www.metaboanalyst.ca), a web-based metabolomics data processing tool [32].
Institutions
- University of Saskatchewan