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Whereas urine and blood are typically targeted in clinical research, saliva represents an interesting alternative because its intrinsic metabolome is chemically diverse and reflective for various biological processes. Moreover, saliva collection is easy and noninvasive, which is especially valuable for cohorts in which sample collection is challenging, for example, infants and children. With this rationale, we established a validated ultra-high-performance liquid chromatography high-resolution mass spectrometry (UHPLC-HRMS) method for salivary metabolic profiling and fingerprinting. Hereby, 450 mu L of saliva was centrifuged and passed over a 0.45-mu m polyamide membrane filter, after which the extract was subjected to chromatographic analysis (HSS T3 column) and QExactive Orbitrap-MS. For the majority of the profiled metabolites, good linearity (R-2 >= 0.99) and precision (coefficient of variance = 0.99). In addition, the method was proven fitfor-purpose for a cohort of 140 adolescents (6-16 years, stratified according to weight), yielding relevant profiles (45 obesity-related metabolites) and discriminative fingerprints (Q(2) of 0.784 for supervised discriminant analysis). Alternatively, laser-assisted rapid evaporative ionization mass spectrometry (LA-REIMS) was established for rapid fingerprinting of saliva, thereby using a Nd:YAG laser and Xevo G2-XS QToF-MS. With an acquisition time of 0.5 min per sample, LA-REIMS offers unique opportunities for point-of-care applications. In conclusion, this work presents a platform of UHPLC-HRMS and LA-REIMS, complementing each other to perform salivary metabolomics.

As lipids are assigned a plethora of biological functions, it is evident that dysregulated lipid metabolism signifies a key element in many pathological conditions. With this rationale, this study presents a validated lipidomics platform to map the fecal lipidome, which integrates unique information about host-gut microbiome interactions, gastrointestinal functionality, and dietary patterns. This particular method accomplished coverage across all eight lipid categories: fatty acyls, glycerolipids, phosphoglycerolipids, polyketides, prenols, saccharolipids, sphingolipids, and sterols. Generic extraction of freeze-dried feces was achieved by solid-liquid extraction using methanol and methyl tert-butyl ether. Extracted components were separated by liquid chromatography, whereby the selected ethylene-bridged hybrid phenyl ultra-high-performance liquid chromatography stationary phase allowed fast separation of both individual lipid species and categories. Detection was achieved by high-resolution full-scan Q-Exactive Orbitrap mass spectrometry and covered a broad m/z scan range (67-2300 Da). Method validation was performed in a targeted fashion to evaluate the analytical performance across all lipid categories, revealing excellent linearity (R