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The natural  15 N/ 14 N isotope composition (δ 15 N) of a tissue is a consequence of its N source and N physiological mechanisms in response to the environment. It could potentially be used as a tracer of N metabolism in plants under changing environmental conditions, where primary N metabolism may be complex, and losses and gains of N fluctuate over time. In order to test the utility of δ 15 N as an indicator of plant N status in N 2 -fixing plants grown under various environmental conditions, alfalfa ( Medicago sativa  L.) plants were subjected to distinct conditions of [CO 2 ] (400 vs. 700 μmol mol −1 ), temperature (ambient vs. ambient +4°C) and water availability (fully watered vs. water deficiency—WD). As expected, increased [CO 2 ] and temperature stimulated photosynthetic rates and plant growth, whereas these parameters were negatively affected by WD. The determination of δ 15 N in leaves, stems, roots, and nodules showed that leaves were the most representative organs of the plant response to increased [CO 2 ] and WD. Depletion of heavier N isotopes in plants grown under higher [CO 2 ] and WD conditions reflected decreased transpiration rates, but could also be related to a higher N demand in leaves, as suggested by the decreased leaf N and total soluble protein (TSP) contents detected at 700 μmol mol −1  [CO 2 ] and WD conditions. In summary, leaf δ 15 N provides relevant information integrating parameters which condition plant responsiveness (e.g., photosynthesis, TSP, N demand, and water transpiration) to environmental conditions.

Leaf δ15N as a physiological indicator of the responsiveness of N2-fixing alfalfa plants to elevated [CO2], temperature and low water availability