This study was undertaken to cope with artifacts derived from CaCO3-CO2-H2O equilibria that occur in substrate-induced respiration (SIR) measurements in calcareous soils. Considering that partitioning of respired CO2 between headspace and soil solution becomes increasingly important above a pH value of 6 and when headspace-to-solution ratios are low, an underestimation in respiration measurements are to be expected under such conditions. In order to evaluate the relative magnitude of this bias in respiration measurements under a wide range of soil conditions, we investigated soils from four sites along a climatic gradient in Israel. Even though all soils were formed over calcareous bedrock, their CaCO3 contents increased with increasing climate aridity. We calculated the theoretical distribution of CO2 between the gaseous and solution phases for each soil, based on the equilibrium reactions between CO2, water, and the various carbonate species. Theoretically calculated factors required for the correction of biased CO2 evolution data were compared to those obtained by actually measuring CO2 retention in the soils, and were found to be smaller, although reasonably close to them. CO2 retention levels determined by experiment were substantial and followed the order: Mediterranean (74% of initial headspace CO2) > semi-arid (70%) > humid-Mediterranean (64%) > arid soil (58%). An additional potential artifact in substrate utilization profiling was addressed when soil respiration was found to be overestimated when induced by considerably acidic substrates such as carboxylic acids. This overestimation results from soil pH lowering which reduces the CO2 retention potential. Hence, if CO2 retention correction factors are applied to carboxylic acid-induced respiration measurements, they must be reduced according to any pH decline. Moreover, CaCO3 dissolution may follow pH decline, and be accompanied by abiotic CO2 evolution that may be mistakenly regarded as respired CO2. We suggest taking the following sequence of measures when generating substrate utilization profiles for calcareous soils: (1) experimental determination of CO2 retention levels and retention correction factors; (2) comparison with theoretical calculations; (3) assessment of pH changes induced by substrates and modification of retention factors necessitated by these changes; (4) measurement of abiotic substrate-induced CO2 evolution rates and their subtraction from measured SIR data; and (5) multiplication of biotic SIR data by CO2 retention correction factors. The comparative analysis of utilization profiles obtained from corrected SIR data with those obtained from the originally measured CO2 evolution rates emphasized the immense underestimation of non-acidic SIRs, concurrent with an overestimation of SIRs obtained for appreciably acidic substrates such as carboxylic acids, in ordinary SIR assays.
Bibliographical noteFunding Information:
We wish to thank Mrs. Ginetta Barness for technical assistance and special thanks to Ms. Sharon Victor for her useful comments. This research is part of the PhD thesis of Adi Oren. This research was supported by the Israel Science Foundation.
- Abiotic CO evolution
- CO retention
- CaCO-CO-HO equilibria
- Data correction
- Substrate utilization profiles