Compound-specific carbon isotope analysis of post-photosynthetic metabolites in C3 and C4 grasses

Israel WK1, Koller F1, Hocart C2, Tcherkez G2, Stuart-Williams H2 and Ghannoum O1

  1. ARC Centre of Excellence for Translational Photosynthesis, Hawkesbury Institute for the Environment, Western Sydney University.
  2. ARC Centre of Excellence for Translational Photosynthesis, The Australian National University.

Breeding crops with improved water-use efficiency (WUEi) has been successful through examining the integrated ratio of the naturally occurring stable carbon isotopes (13C /12C or δ13 C) incorporated during photosynthesis. It is intensively studied among C3 species most notably that of wheat varieties with improved WUE. The carbon concentrating mechanism (CCM) in C4 allows for an enriched δ13C signature relative to C3. Since C4 species are known to have better WUE, attempts have been undertaken to correlate δ13 C with WUE i. In C3 species, WUE is directly correlated to Ci/Ca which can be measured by analysing δ13C. However, δ13C and thus WUE in C4 plants are not consistently correlated because of the CCM and other leaf metabolic reactions including post-photosynthetic fractionation. Thus, the study seeks to examine δ13C signatures of post-photosynthetic metabolites among C4 grasses to ultimately understand the mechanisms of intrinsic water use efficiency; hence, providing a breeding tool for crops with improved WUE. Grasses representing C3 and C4 photosynthetic types and C4 biochemical subtypes (NADP-ME, NAD-ME, and PCK) were grown in glasshouse conditions and whole leaves were assayed for simple sugar, organic acids, and bulk metabolic pool extraction and purification. δ13C signatures were assayed using HPLC coupled with wet oxidiser that directly converts eluted compounds to CO2 and measured using IRMS. Results show that fructose is 13C-enriched by 2.8 ‰ in NADP-ME relative to NAD-ME while sucrose and glucose are not significantly different. PCK metabolites tend to have an intermediate 13C signature when compared to NADP-ME and NAD-ME while the greatest depletion in 13C was observed among C3. These results indicate initial evidence that could potentially explain the variations in 13C signatures among C4 grasses and ultimately develop screening tools for improved WUE.