SCFAs were extracted and analyzed as described previously (49). An NMR spectrum for each fecal sample was acquired using NOESY-presaturation on a 4-channel Bruker Ascend 700 MHz spectrometer (Bruker Biospin) with a TXI 700 MHz 3 mm probe with triple-axis gradients. All spectra were acquired with a 14 ppm sweep width, 4-second acquisition time, 4 dummy scans, and 384 transients. All spectra were zero filled to 128,000 data points, Fourier transformed with 0.1 Hz line broadening applied, manually phased, and baseline-corrected using TopSpin NMR software. The SCFA concentrations were quantitatively determined using the 700 MHz library from Chenomx NMR Suite 7.1 after acquiring an NMR spectrum from each fecal sample. The resulting quantified metabolites were used as input variables into subsequent statistical analysis and normalized by fecal weight.

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