Does metabolic water control the phosphate oxygen isotopes of microbial cells?

The oxygen isotopes ratio (δ18O) of microbial cell water strongly controls the δ18O of cell phosphate and of other oxygen-carrying moieties. Recently it was suggested that the isotopic ratio in cell water is controlled by metabolic water, which is the water produced by cellular respiration. This potentially has important implications for paleoclimate reconstruction, and for measuring microbial carbon use efficiency with the 18O-water method. Carbon use efficiency strongly controls soil organic matter preservation. Here, we directly tested the effect of metabolic water on microbial cells, by conducting experiments with varying the δ18O of headspace O2 and the medium water, and by measuring the δ18O of cell phosphate. The latter is usually assumed to be in isotopic equilibrium with the cell's water. Our results showed no correlation between the δ18O of O2 and that of the cell phosphate, contradicting the hypothesis that metabolic water is an important driver of δ18O of microbial cell water. However, our labeled 18O water experiments indicated that only 43% of the oxygen in the cell's phosphate is derived from equilibration with the medium water, during late-log to early-stationary growing phase. This could be explained by the isotopic effects of intra-and extra-cellular hydrolysis of organic compounds containing phosphate.

Soil phosphate stable oxygen isotopes across rainfall and bedrock gradients.

The stable oxygen isotope compositions of soil phosphate (δ(18)O(p)) were suggested recently to be a tracer of phosphorus cycling in soils and plants. Here we present a survey of bioavailable (resin-extractable or resin-P) inorganic phosphate δ(18)O(p) across natural and experimental rainfall gradients, and across soil formed on sedimentary and igneous bedrock. In addition, we analyzed the soil HCl-extractable inorganic δ(18)O(p), which mainly represents calcium-bound inorganic phosphate. The resin-P values were in the range 14.5-21.2‰. A similar range, 15.6-21.3‰, was found for the HCl-extractable inorganic δ(18)O(p), with the exception of samples from a soil of igneous origin that show lower values, 8.2-10.9‰, which indicate that a large fraction of the inorganic phosphate in this soil is still in the form of a primary mineral. The available-P δ(18)O(p) values are considerably higher than the values we calculated for extracellular hydrolysis of organic phosphate, based on the known fractionation from lab experiments. However, these values are close to the values expected for enzymatic-mediated phosphate equilibration with soil-water. The possible processes that can explain this observation are (1) extracellular equilibration of the inorganic phosphate in the soil; (2) fractionations in the soil are different than the ones measured at the lab; (3) effect of fractionation during uptake; and (4) a flux of intercellular-equilibrated inorganic phosphate from the soil microbiota, which is considerably larger than the flux of hydrolyzed organic-P.

A method for analyzing the δ18O of resin-extractable soil inorganic phosphate.

Improved tools for tracing phosphate transformations in soils are much needed, and can lead to a better understanding of the terrestrial phosphorus cycle. The oxygen stable isotopes in soil phosphate are still not exploited in this regard. Here we present a method for measuring the oxygen stable isotopes in a fraction of the soil phosphate which is rapidly available to plants, the resin-extractable P. This method is based on extracting available phosphate from the soil with anion-exchange membranes, soil organic matter removal by a resin, purification by precipitation as cerium phosphate, and finally precipitation as silver phosphate. The purified silver phosphate samples are then measured by a high-temperature elemental analyzer (HT-EA) coupled in continuous flow mode to an isotope ratio mass spectrometer. Testing the method with Mediterranean and semi-arid soils showed no artifacts, as well as good reproducibility in the same order as that of the HT-EA analytical uncertainty (0.3‰).