Mass profiler professional software

Experiments were performed in triplicate, and the results were expressed as mean ± standard error of the mean (SEM). Data were analyzed through one-way analysis of variance (ANOVA) in conjunction with Duncan’s multiple range test using SPSS software (version 26.0, SPSS, Inc., Chicago, IL, USA). The p-value was set to 0.05 for a significant difference.
The MS spectra were analyzed and converted to compound Exchange Format (.CEF) files with the help of the “find compounds by molecular feature” tool using Agilent Mass Hunter Qualitative Analysis Software (version B.07.00). The exported files were then imported into Mass Profiler Professional (MPP) software (version 14.0, Agilent Technologies, Santa Clara, CA, USA) for further statistical analysis. Alignment parameters were: RT window = 0.5% + 0.1 min, mass window = 10 ppm + 1 mDa. A principal component analysis (PCA) was performed with MPP to visualize the sample groupings at the different stages of in vitro simulated digestion. The resulting entity list was then processed in the ID browser, which allowed for chemical formulas to be generated and searched against a proprietary database.

An acetonitrile/methanol (1:1, v/v) solution was incorporated into serum samples to obtain the metabolites. The mixture was vortexed for 30 seconds, sonicated at 4°C for 10 minutes and then left at -80°C overnight. On the second day, the samples were centrifuged at 12,000 rpm at 4°C for 15 minutes. The supernatant was debrided, and the pellet was desiccated after vacuum centrifugation. The samples were redissolved in acetonitrile/methanol mixture (150 μL, 1:1, v/v), sonicated at 4°C for 10 minutes, centrifuged at 12,000 rpm for 15 minutes and incubated at -80°C overnight. On the third day the supernatants were dissolved at 4°C and filtered through a nylon membrane (0.22 µm) into sample bottles for LC-MS/MS analysis.
Mass spectrometry conditions: capillary voltage: 3.5 kV positive ion, 3.5 kV negative ion, ion source temp: 325°C, the flow rate of drying gas: 10 L/min, atomization pressure: 35 psi, sheath temp: 370°C, gas flow rate in sheath: 12 L/min.
Raw data were processed using Agilent Profinder software to carry out retention time correction and identification, extraction, integration and alignment of peaks, etc., finally output CEF file. Statistical analysis was conducted using the Agilent Mass Profiler Professional (MPP) software, the Metlin database was also utilized for the identification of substances together with metabolic pathway analysis.