NMR Spectroscopy of Biological Samples

Urine, feces, serum, and tissue samples were prepared according to previously published protocols [27] (link), [35] (link). Individual BALF samples (200 µl) were mixed with phosphate buffer (400 µl) and 550 µl of the mixture was subsequently transferred into 5 mm diameter standard NMR tubes for data acquisition (Bruker Biospin, Rheinstetten). All biological samples were analyzed using a 600 MHz Avance DRX NMR spectrometer (Bruker; Rheinstetten), employing a standard one dimensional (1D) ¹H NMR Noesypr1d pulse sequence with water suppression (recycle delay (rd)-90°-t1-90°-tm-90°-acquisition time) for all samples. Recycle delay (rd) and mixing time (tm) were set to 2 s and 100 ms respectively. For obtaining more comprehensive information on the serum samples, two additional pulse programs were utilized namely Carr-Purcell-Meiboom-Gill (cpmgpr) and diffusion edited spectroscopy (ledbipolpr) [35] (link). The numbers of scans were adjusted for each biological matrix, whereby urine and serum were analyzed in 256 and extracts in 128 scans to obtain maximum signal output. Data from each sample was accumulated into 32 k data points within a spectral width of 20 ppm.