Spectra were collected on a 700 MHz Bruker Avance III HD spectrometer (Bruker, Milton, ON, Canada). The Bruker 1-D NOESY gradient water suppression pulse sequence ‘noesygppr1d’ was used with 10 ms mixing time. Each sample was run for 512 scans to a total acquisition size of 128 k, a spectral window of 20.5 ppm, a transmitter offset of 4.7 ppm, and a recycle delay of 4 s. All measurements were recorded using a Bruker triple resonance TBO-Z probe. The Bruker automation program “pulsal” was used on each sample before data acquisition to guarantee that the 90-degree pulse was calibrated correctly, ensuring quantitative and comparable data across samples [26 (link)]. The spectra were zero filled to 256 k, automatically phased, baseline corrected, and line-broadened by 0.3 Hz [25 (link)]. Spectra were then exported to MATLAB (MathWorks, Natick, MA, USA) as ASCII files, and underwent dynamic adaptive binning [30 (link)], followed by manual inspection and correction. Spectral binning resulted in 439, 460, and 379 spectral bins for kidney, liver, and breast muscle, respectively. The dataset was then normalized to the total metabolome, excluding the region containing the water peak, and pareto scaled.
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