Salvadora persica L. (toothbrush tree, Miswak) is well recognized in most Middle Eastern and African
countries for its potential role in dental care, albeit the underlying mechanism for its effectiveness is still
not fully understood. A comparative MS and NMR metabolomics approach was employed to investigate
the major primary and secondary metabolites composition of S. persica in context of its organ type viz.,
root or stem to rationalize for its use as a tooth brush. NMR metabolomics revealed its enrichment in
nitrogenous compounds including proline-betaines i.e., 4-hydroxy-stachydrine and stachydrine reported
for the first time in S. persica. LC/MS metabolomics identified flavonoids (8), benzylurea derivatives (5),
butanediamides (3), phenolic acids (8) and 5 sulfur compounds, with 21 constituents reported for the
first time in S. persica. Principal component analysis (PCA) and hierarchical cluster analysis (HCA) of
either NMR or LC/MS dataset clearly separated stem from root specimens based on nitrogenous compounds abundance in roots and is justifying for its preference as toothbrush versus stems. The presence
of betaines at high levels in S. persica (9−12 g/mg dry weight) offers novel insights into its functioning
as an osmoprotectant that maintains the hydration of oral mucosa. Additionally, the previously described
anti-inflammatory activity of stachydrine along with the antimicrobial effects of sulfonated flavonoids,
benzylisothiocynate and ellagic acid derivatives are likely contributors to S. persica oral hygiene health
benefits. Among root samples, variation in sugars and organic acids levels were the main discriminatory criterion. This study provides the first standardization of S. persica extract using qNMR for further
inclusion in nutraceuticals.