Investigating Metabolomics Analytical Techniques to Analyze and Better Understand the Human Saliva Metabolome

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The Ohio State University

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As one of the most rapidly evolving fields in biomedical research, metabolomics offers small molecule compositional analysis of biological fluids, cells, and tissues. The metabolome itself can provide insights into biological mechanisms of disease. Chemical phenotyping using biospecimens and metabolomics can lead to the identification of biomarkers which may underlie diseases. Saliva, in particular, contains multiple biomarkers making it suitable for metabolomics. Despite the fact that saliva testing already exists and offers a non-invasive and rapid method of analysis, characterization of the human saliva metabolome and saliva-related metabolomics applications are not very extensive. Additionally, current nuclear magnetic resonance (NMR) saliva analysis often involves several time-consuming steps that increase the time of analysis and may have a negative impact on the analysis. This project aims to investigate more suitable metabolomic analysis techniques of NMR on saliva. A comparison between pulse sequences (CPMG and 1D NOESY) and magnetic fields (600 and 700 MHz), as well as evaluation of sample storage stability was conducted with 1:1 saliva, sodium phosphate buffer solution samples. Results indicated that metabolite peak shifts were not significantly affected by NMR sequences and instruments but the water peak suppression in CPMG pulse sequencing with 700 MHz magnetic field, deemed most efficient. In addition, the CPMG experiment produces spectra with slightly better baseline in some cases. Sample stability persisted in spectra analysis between tested samples based on storage conditions of 1 week in the freezer and 24 hours at room temperature. After method optimization, NMR spectra were collected and used to assess the influence of genetic and environmental factors on the saliva metabolome. More specifically, 119 saliva samples from smokers and non-smokers were analyzed in total. Finally, a systematic 2D NMR study was conducted to perform the unambiguous assignment of the NMR spectra of saliva. This is important for performing biomarker identification and pathway analysis which can further contribute to foods impact on the saliva metabolome.



Saliva, Metabolomics, NMR