Bicarbonate as tracer for assimilated C and homogeneity of ¹⁴C and ¹⁵N distribution in plants by alternative labeling approaches

Abstract

Aims

Application of carbon (C) and nitrogen (N) isotopes is an essential tool to study C and N flows in plant-soil-microorganisms systems. When targeting single plants in a community the tracers need to be added via e.g., leaf-labeling or stem-feeding approaches. In this study we: (i) investigated if bicarbonate can be used to introduce ¹⁴C (or ¹³C) into white clover and ryegrass, and (ii) compared the patterns of ¹⁴C and ¹⁵N allocation in white clover and ryegrass to evaluate the homogeneity of tracer distribution after two alternative labeling approaches.

Methods

Perennial ryegrass and white clover were pulse labeled with ¹⁵N urea via leaf-labeling and ¹⁴C either via a ¹⁴CO₂ atm or with ¹⁴C bicarbonate through leaf-labeling. Plants were sampled 4 days after labeling and prepared for bulk isotope analysis and for ¹⁴C imaging to identify plant parts with high and low ¹⁴C activity. Subsequently, plant parts with high and low ¹⁴C activity were separated and analyzed for ¹⁵N enrichment.

Results

Bicarbonate applied by leaf-labeling efficiently introduced ¹⁴C into both white clover and ryegrass, although the ¹⁴C activity in particular for white clover was found predominantly in the labeled leaf. Using ¹⁴C imaging for identification of areas with high (hotspots) and low ¹⁴C activity showed that ¹⁴C was incorporated very heterogeneously both when using bicarbonate and CO₂ as expected when using pulse labeling. Subsequent analysis of ¹⁵N enrichment in plant parts with high and low ¹⁴C activity showed that ¹⁵N also had a heterogeneous distribution (up to two orders of magnitude).

Conclusion

Bicarbonate can efficiently be used to introduce ¹⁴C or ¹³C into plant via the leaf-labeling method. Both ¹⁴C and ¹⁵N showed heterogeneous distribution in the plant, although the distribution of ¹⁵N was more even than that of ¹⁴C.