Isotopic Criteria in the Characterization of Aromatic Molecules. 1. Hydrogen Affiliation in Natural Benzenoid/Phenylpropanoid Molecules

Abstract

The site-specific isotope ratios of several families of aromatic molecules are analyzed in terms of hydrogen affiliation and discriminating potential. Among the aromatic molecules produced by plants, many are biosynthesized by the shikimate pathway, but the terpenic pathway also forms some compounds with a benzenic ring. In compounds of the phenylpropanoid family, specific hydrogen connections are determined with cinnamic acid, a key intermediate in the formation of a large number of aromatic molecules. Then affiliations through the phenylalanine precursor, back to the parent d-erythrose 4-phosphate and phosphoenolpyruvate molecules and finally to glucose, are considered. Typical isotopic profiles of the benzenic ring in natural, as compared to non-natural, molecules are defined. The dispersion observed in the (D/H)i ratios of the lateral chains is illustrative of diverse mechanistic responses and the role of exchange phenomena. The isotopic patterns of aromatic molecules pertaining to the terpenic family are drastically different from those of the shikimate descendants, and they exhibit much less variability. They enable the stereochemical affiliation of individual hydrogen atoms to be traced back first to the parent atoms in the common intermediate, geranyl diphosphate, then to the glyceraldehyde 3-phosphate and pyruvate couple involved in the DOXP pathway, and ultimately to the glucose precursor. The results illustrate the aptitude of the site-specific isotope ratios not only to authenticate natural with respect to chemical molecules but also to characterize different metabolic pathways and to reveal differences associated with the nature of the plant precursor.

Multi-factorialin vivostable isotope fractionation: causes, correlations, consequences and applications

“…The relative positional 2 H abundance or ratio of isotopomers is p > o ≥ m for natural aromatic compounds and can be deduced from that of the precursors and from intramolecular (in between isotopomer molecules occurring) KIEs on defined reactions. A hydroxylation, for example, at the p-position, implies the so-called NIH-shift, shifting the relative 2 H enrichment into the neighboured mpositions and changing the 2 H pattern of the non-substituted positions to m > o [33,206,209]. Application of knowledge about the involved reaction mechanism and implied KIEs allowed the prediction of 2 H patterns of the products of o-hydroxylation.…”

Analytical contribution of deuterium 2D‐NMR in oriented media to 2H/1H isotopic characterization: the case of vanillin

“…Martin et al . reported the mean 2 H/ 1 H ratio values for the C 3 and C 5 positions of a range of 4‐hydroxylated natural aromatic compounds . In all cases these positions are enriched relative to that also reported for the C4 of benzaldehyde and methyl cinnamate; however, such interpretations need to be treated with caution.…”

“…In all cases these positions are enriched relative to that also reported for the C4 of benzaldehyde and methyl cinnamate; however, such interpretations need to be treated with caution. As detailed in this paper, considerable variation in the values for precursors, variation in pathways, exchange reactions during biosynthesis, or during extraction and purification, and uncertainties such as the extent to which the NAD(P)H pool is depleted relative to cell water can all act to introduce ‘noise’ into the processing.…”

Isotopic Criteria in the Characterization of Aromatic Molecules. 2. Influence of the Chemical Elaboration Process

“…In a context where the natural status of products is considered as an important quality criterion, it is of prime importance to be able to ascertain, for a given molecule, whether it has been extracted from plants or has been obtained by chemical synthesis using either fossil or natural raw materials. We have shown, in parallel (2), that isotopic distributions are a rich source of information on the biogenesis of natural aromatic products. In comparison, Table 2 gives the basis for easily differentiating plant molecules from their synthetic counterpart and illustrates the aptitude of isotopic parameters for providing further information on the chemical history of the molecule.…”

“…In particular, site-specific hydrogen isotope ratios measured by NMR (SNIF-NMR), possibly combined with overall carbon or oxygen isotope parameters determined by isotope ratio mass spectrometry (IRMS), are now currently used to authenticate a number of natural aroma components (1). In this context, we have investigated, in parallel, the isotopic affiliation in benzenoid/phenylpropanoid molecules biosynthesized by plants (2).…”