Estimation of Nitrogen Fixation by Isotope Dilution in Field and Greenhouse Experiments1

Abstract

Isotope dilution methodology offers the advantage of an integrative technique for measuring N₂ fixation in the field, but may have limitations in grass‐legume mixtures because of the possibility of transfer of fixed N from legume to grass. A study was conducted in the greenhouse and in the field utilizing two soils, Hanford sandy loam and Yolo silt loam (Typic Xerorthents) in which the organic matter was labeled by previous application of either ¹⁵N‐depleted or ¹⁵N‐enriched compounds. Contributions of atmospheric N₂ to total plant N were calculated from the isotopic composition of plant tissue, using the composition of the non‐legume as a reference value.Ladino clover (Trifolium repens L.) and Wimmera ryegrass (Lolium rigidum L.) were grown alone and in mixture for purposes of comparison. In the greenhouse estimation of N₂ fixation by the isotope and difference methods were in good agreement. There was essentially no transfer of fixed N₂ from the clover to the ryegrass in any of the pot experiments. In the field experiment, clover obtained 85 to 100% of its N from the fixation process. In the clover‐ryegrass mixture, % total N and % ¹⁵N excess comparisons both indicated substantial transfer of N from clover to ryegrass after the stand had been established after about 6 months. Up to 79% of the N in ryegrass was calculated to be derived from such transfer. It is concluded that the isotope dilution method gives estimates of N₂ fixation at least as good as those obtained by yield‐dependent methods such as the difference method, but it is not suitable for grass‐legume mixtures.