Saliva NMR metabolomics: Analytical issues in pediatric oral health research

Abstract

Objectives Saliva metabolome is a promising diagnostic tool concerning oral and systemic diseases. We aimed at establishing a suitable protocol for saliva collection and gauging the relative impacts of gender, dentition stage, and caries on the saliva metabolome of a small children cohort. Subjects and methods A nuclear magnetic resonance‐based metabolomics cross‐sectional study of children saliva (n = 38) compared the effects of: (a) stimulation and unstimulation conditions, and (b) collection through passive drool and using an absorbing device. Multivariate and univariate statistical analyses were applied to evaluate such effects and those related to gender, dentition stage and caries. Results No significant differences were found between unstimulated and stimulated saliva, and the former was used for subsequent studies. Swab collection induced significant changes in sample composition, indicating passive drool as preferential. The impacts of gender and dentition stage were not significant compared to that of caries, which induced variations in the levels of 21 metabolites. These comprised amino acids and monosaccharides observed for the first time to our knowledge regarding children caries, suggesting protein hydrolysis and deglycosylation. Conclusions Unstimulated passive drool saliva metabolome may carry a caries signature.

Targeted metabolomics of pellicle and saliva in children with different caries activity

“…Lactate and 1-methylhistine are two of the metabolites that emerged in the literature previously in the context of caries activity 31,32 . However, data are conflicting, as increased lactate concentrations in saliva of caries-active children have been reported 32 but not confirmed 33 . In vitro experiments revealed that lactate exposition leads to tooth enamel demineralisation to a greater extent than acetate or propionate 34 .…”

“…L-serine, L-pyroglutamate and L-threonine showed lower concentrations in the inactive group than in the other two groups, whereas it was opposite for the other two compounds. For nine other amino acids (alanine, aspartate, glutamine, glycine, isoleucine, leucine, proline, taurine, and tyrosine), an increased concentration in swap-collected unstimulated saliva of caries-active children has been shown 33 . However, our data did not confirm this.…”

“…Furthermore, after a longer incubation time in the oral cavity, differences due to bacterial adhesion could possibly be detected, e.g. an accumulation of organic acids produced by microorganisms or altered concentrations of carbohydrates due to the hydrolytic cleavage of glycoproteins by bacterial enzymes 33 . However, from our experience, incubation times of more than 10 min for pellicle collection within a study of children ages ≤6 years is hard to realise.…”

Salivary Metabolomics: From Diagnostic Biomarker Discovery to Investigating Biological Function

“…Furthermore, some studies have reported conflicting results. For example, butyrate has been found to be increase or decrease in concentration in association with the presence of dental caries [45,46]. It is likely that single metabolites in isolation may not yield sufficient diagnostic specificity for most disease, however multivariate analyses may offer improved accuracy.…”

“…1 H-NMR analyses have found salivary metabolite profile is independent of sex and BMI when considering net metabolite profile [86], although sex-based metabolic differences have been found when quantifying individual metabolites [91]. Collection method is another caveat for the metabolomic study of saliva, with the use of collection systems as opposed to passive drooling being identified as a potential source of exogenous analytes in samples [46].…”