O of leaf water, d18 O-PO 4 , soybean, structural P, TCA-soluble reactive P (TCA P). SummaryThe objective of this study was to investigate the isotopic composition of oxygen bound to phosphate (d 18 O-PO 4 ) in different phosphorus (P) pools in plant leaves. As a model plant we used soybean (Glycine max cv Toliman) grown in the presence of ample P in hydroponic cultures.The leaf blades were extracted with 0.3 M trichloroacetic acid (TCA) and with 10 M nitric acid. These extractions allowed measurement of the TCA-soluble reactive P (TCA P) that is rapidly cycled within the cell and the total leaf P. The difference between total leaf P and TCA P yielded the structural P which includes organic P compounds not extractable by TCA.P
Organic fertilizer applications can contribute to Zinc (Zn) biofortification of crops. An enriched stable isotope source tracing approach is a central tool to further determine the potential of this biofortification measure. Here, we assessed the use of the widely available quadrupole single-collector ICPMS (Q-ICPMS, analytical error = 1% relative standard deviation) and the less accessible but more precise multicollector ICPMS as reference instrument (MC-ICPMS, analytical error = 0.01% relative standard deviation) to measure enriched Zn stable isotope ratios in soil–fertilizer–plant systems. The isotope label was either applied to the fertilizer (direct method) or to the soil available Zn pool that was determined by isotope ratios measurements of the shoots that grew on labeled soils without fertilizer addition (indirect method). The latter approach is used to trace Zn that was added to soils with complex insoluble organic fertilizers that are difficult to label homogeneously. To reduce isobaric interferences during Zn isotope measurements, ion exchange chromatography was used to separate the Zn from the sample matrix. The 67Zn:66Zn isotope ratios altered from 0.148 at natural abundance to 1.561 in the fertilizer of the direct method and 0.218 to 0.305 in soil available Zn of the indirect method. Analysis of the difference (Bland–Altman) between the two analytical instruments revealed that the variation between 67Zn:66Zn isotope ratios measured with Q-ICPMS and MC-ICPMS were on average 0.08% [95% confidence interval (CI) = 0.68%]. The fractions of Zn derived from the fertilizer in the plant were on average 0.16% higher (CI = 0.49%) when analyzed with Q- compared to MC-ICPMS. The sample matrix had a larger impact on isotope measurements than the choice of analytical instrument, as non-purified samples resulted on average 5.79% (CI = 9.47%) higher isotope ratios than purified samples. Furthermore, the gain in analytical precision using MC-ICPMS instead of Q-ICPMS was small compared to the experimental precision. Thus, Zn isotope measurements of purified samples measured with Q-ICPMS is a valid method to trace Zn sources in soil–fertilizer–plant systems. For the indirect source tracing approach, we outlined strategies to sufficiently enrich the soil with Zn isotopes without significantly altering the soil available Zn pool.
Manure and sewage sludge are known to add significant amounts of zinc (Zn) and other metals to soils. However, there is a paucity of information on the fate of Zn that derives from complex organic fertilizers in soil–plant systems and the contribution of these fertilizers to the Zn nutrition of crops. To answer these questions, we grew Italian ryegrass in the presence of ZnSO4, sewage sludge, and cattle and poultry manure in an acidic soil from Heitenried, Switzerland, and an alkaline soil from Strickhof, Switzerland, where the isotopically exchangeable Zn had been labeled with 67Zn. This allowed us to calculate the fraction of Zn in the shoots that was derived from fertilizer, soil, and seed over 4 successive cuts. In addition, we measured the 67Zn:66Zn isotope ratio with the diffusive gradients in thin films technique (DGT) on soils labeled with 67Zn and incubated with the same fertilizers. After 48 days of growth, the largest fraction of Zn in the ryegrass shoots was derived from the soil (79–88%), followed by the Zn-containing fertilizer (11–20%); the least (<2.3%) came from the seed. Only a minor fraction of the Zn applied with the fertilizer was transferred to the shoots (4.7–12%), which indicates that most of the freshly added Zn remained in the soil after one crop cycle and may thereby contribute to a residual Zn pool in the soil. The 67Zn:66Zn isotope ratios in the DGT extracts and the shoots measured at cut 4 were identical, suggesting that the DGT and plant took up Zn from the same pool. The proportion of Zn derived from the fertilizers in the DGT extracts was also identical to that measured in ryegrass shoots at cut 4. In conclusion, this work shows that stable Zn isotope labeling of the soil available Zn can be used to precisely quantify the impact of complex organic fertilizers on the Zn nutrition of crops. It also demonstrates that DGT extractions on labeled soils could be used to estimate the contribution of Zn fertilizers to plant nutrition.
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