Metabolomic profile
CPMG-BRUKER
1H nuclear magnetic resonance (NMR) was used to analyze the metabolomic profile. It was carried out at the Multiuser Laboratory of Characterization and Analysis (LMCA) at UFPB in a Bruker model spectrometer operating at 400 MHz. The pulse sequence used to obtain the 1H spectra was CPMG PR1D, number of scans: 128, receiver gain: 58.8, acquisition time: 4.08s, spectral size: 64k and temperature: 30 °C and were performed in triplicates.Initially, 5 ml of blood was collected in polypropylene tubes with EDTA in the early morning, during fasting. After collection, it was centrifuged at 3000 xg for 10 min at 4 °C to isolate the serum. Serum was transferred to a sterilized tube and stored at − 80 °C. It was thawed for use. 300 µL of serum was collected and 300 µL of phosphate buffer was added as a reference standard. Vortex homogenization was performed for 30 s, immediately after centrifiguation at 14000 x g for 10 min at 4 °C. After centrifugation, 60 µL of deuterated water was added. Then, analysis was performed in the NMR apparatus. The supernatant was transferred to a 400 MHz NMR tube in a 5 mm tube. The pulse sequence used to obtain data was CPMG-BRUKER and analyses were performed in triplicate.The obtained spectra were analyzed using the TopSpin 4.1.1 software and chemical shifts were expressed in ppm. Metabolites were identified by comparing chemical shifts with chemical shifts of authentic samples available in the Human Metabolome Database (HMDB). Spectra were referenced by tetramethylsilane (TMS), whose total range chosen for analysis was between 0.1 and 9 ppm, with binnings of 0.02 ppm. This generated a data matrix of 96 samples and 225 variables, which was exported in the ASCII format for the “The Unscrumbler” statistics software (version 9.7, CAMO Process). Data were not normalized by area, and peaks referring to TMS and water were discarded. With these procedures, it was possible to differentiate phases before and after intervention with mixed dietary fiber.