Saliva as a Complementary Matrix to Whole Blood in Untargeted Lipidomics and Metabolomics: A Baseline Profiling Study in Healthy Subjects

<jats:p>Background/Objectives: Oral fluid (OF) is a promising non-invasive matrix for metabolomic and lipidomic profiling, owing to its simple and repeatable collection. However, a direct, simultaneous comparison of the OF and whole blood (WB) lipidome and metabolome using a harmonized analytical workflow in healthy subjects has not yet been systematically addressed. This study aimed to characterize and compare the untargeted lipidomic and metabolomic profiles of OF and WB in a cohort of healthy adults, and to identify the degree of biochemical overlap and matrix-specific molecular signatures between the two biofluids. Methods: Untargeted HPLC/TOF-MS-based lipidomic and metabolomic analyses were performed on OF (n = 16) and WB (n = 21) samples collected simultaneously from 21 healthy subjects (mean age 29.6 ± 4 years). Data were processed using interquartile range filtering (IQR = 10%), logarithmic transformation, and autoscaling. Multivariate statistical analysis included partial least squares discriminant analysis (PLS-DA), VIP score ranking, hierarchical clustering, and volcano plot analysis (p &amp;lt; 0.05, |log₂FC| &amp;gt; 1). Results: OF yielded 58 metabolites (25 chemical classes) and 730 lipid species (77 subclasses), with amino acids, purines, and ether phosphatidylethanolamines as dominant groups. WB yielded 106 metabolites (24 classes) and 859 lipid species (61 subclasses), with fatty acids, glycerophospholipids, and sphingomyelins predominating. After quality filtering, 262 compounds were shared between the two matrices, with glycerophospholipids, triglycerides, and fatty acids constituting the molecular backbone of their biochemical overlap. PLS-DA revealed a clear matrix-specific separation (Component 1 = 56.4% variance), driven by niacinamide, 2-piperidone, ST 27:1;O;S, and linoleic acid (FA 18:2), whose differential abundance reflects the independent metabolic contribution of the oral cavity. Conclusions: OF and WB generate distinct yet partially overlapping molecular signatures. These findings support the concept that OF represents a complementary biological matrix rather than a simple surrogate of WB and establish a reference lipidomic and metabolomic dataset for the future application of salivary profiling as a non-invasive diagnostic tool.</jats:p>