Quantitative analysis of blood plasma metabolites using isotope enhanced NMR methods.

ABSTRACT: NMR spectroscopy is a powerful analytical tool for both qualitative and quantitative analysis. However, accurate quantitative analysis in complex fluids such as human blood plasma is challenging, and analysis using one-dimensional NMR is limited by signal overlap. It is impractical to use heteronuclear experiments involving natural abundance (13)C on a routine basis due to low sensitivity, despite their improved resolution. Focusing on circumventing such bottlenecks, this study demonstrates the utility of a combination of isotope enhanced NMR experiments to analyze metabolites in human blood plasma. (1)H-(15)N HSQC and (1)H-(13)C HSQC experiments on the isotope tagged samples combined with the conventional (1)H one-dimensional and (1)H-(1)H TOCSY experiments provide quantitative information on a large number of metabolites in plasma. The methods were first tested on a mixture of 28 synthetic analogues of metabolites commonly present in human blood; 27 metabolites in a standard NIST (National Institute of Standards and Technology) human blood plasma were then identified and quantified with an average coefficient of variation of 2.4% for 17 metabolites and 5.6% when all the metabolites were considered. Carboxylic acids and amines represent a majority of the metabolites in body fluids, and their analysis by isotope tagging enables a significant enhancement of the metabolic pool for biomarker discovery applications. Improved sensitivity and resolution of NMR experiments imparted by (15)N and (13)C isotope tagging are attractive for both the enhancement of the detectable metabolic pool and accurate analysis of plasma metabolites. The approach can be easily extended to many additional metabolites in almost any biological mixture.

SUBMITTER: Gowda GA 

PROVIDER: S-EPMC3065653 | biostudies-literature | 2010 Nov

REPOSITORIES: biostudies-literature