A repository of the salivary metabolome and its key drivers in 1436 European children.

BACKGROUND

Saliva is an emerging but understudied biofluid in metabolomics, offering a similar window into biological information as blood, while enabling repetitive and non-invasive sampling.

METHODS

In this observational study, we comprehensively analysed the salivary metabolome of 965 children and adolescents (7-17y) from two exploratory cohorts in Europe, using ultra-high performance liquid chromatography coupled to high-resolution mass spectrometry. Preprocessed data were assessed via (partial) Spearman correlation, Mummichog-based pathway enrichment, and univariate (Welsh t-test, one-way ANOVA, Wilcoxon or Kruskal-Wallis rank-sum test) analyses. All p-values were corrected using the Benjamini-Hochberg procedure. Associations with age, anthropometrics, mental wellbeing, lifestyle, dietary intake and microbiome were explored. Key metabolite associations were validated in two independent cohorts (8-18y, n = 471), yielding a total study population of 1436 participants.

FINDINGS: Over 3500 untargeted features and 188 targeted metabolites were consistently detected in the pooled quality control saliva of the exploratory cohorts. Weight status (|ρ| < 0.255) and age (|ρ| < 0.341) reflected the salivary metabolome best, compared to mental wellbeing (|ρ| < 0.227) and lifestyle factors (|ρ| < 0.185). Multiple metabolites, especially (acetylated) amino acids and derivatives were elevated in participants with overweight or obesity and those experiencing high psychological stress, mirroring blood-based studies. In addition, these metabolites correlated with Bacteroidota, Pseudomonadota, Actinomycetota and Bacillota (|ρ| < 0.423) phyla, suggesting oral microbiome disturbances. Notably, the artificial sweetener acesulfame K, detected for the first time in children and adolescents' saliva, was associated with excess weight (|ρ| < 0.243), psychological stress (|ρ| < 0.148), specific food and/or beverage consumption (|ρ| < 0.348) and Bacillota spp. (|ρ| < 0.196).

INTERPRETATION: Our findings reinforce and expand upon previous blood-based studies, while revealing novel salivary biomarkers and, as such, provide a high-quality repository of the paediatric salivary metabolome for future studies exploring its association with children and adolescent's health.

FUNDING: FAME and OPERA were supported by the Research Foundation Flanders, Ghent University, and the European Research Council. PANIC was funded by Finnish national agencies and foundations. Fin-HIT was funded by the Folkhälsan Research Foundation and the Päivikki and Sakari Sohlberg Foundation.