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DIVISION S-8 - NOTES

Acid fumigation of soils to remove carbonates prior to total organic carbon or CARBON-13 isotopic analysis

^a Stable Isotope Lab., Univ. of California-Davis, Davis, CA 95616
^b Dep. of Land, Air, and Water Resources, Univ. of California-Davis, Davis, CA 95616
^c Dep. of Agronomy and Range Science, Univ. of California-Davis, Davis, CA 95616

* Corresponding author (cvankessel{at}ucdavis.edu)

	ABSTRACT

TOP ABSTRACT INTRODUCTION Materials and Methods Results and Discussion Conclusions REFERENCES

 
The use of ¹³C natural abundance (¹³C) to follow C input to soil has gained widespread acceptance. However, inorganic C present in the soil as carbonates will interfere with the measurement of soil organic ¹³C unless removed or excluded from measurement. We report a simple and convenient HCl-fumigation method to remove inorganic C from soil. Soil samples are weighed in Ag-foil capsules, arranged on a microtiter plate, wetted with water to approximately field capacity, and placed in a desiccator containing a beaker with concentrated (12 M) HCl. The carbonates are released as CO₂ by the acid treatment in 6 to 8 h. The soil samples are then dried at 60°C prior to isotope determination. The advantages of the HCl-fumigation method to remove inorganic C from the soil are that: (i) no water soluble C will be lost from the soil; (ii) a large number of samples can be processed simultaneously; (iii) the removal of inorganic C is rapid and complete; and (iv) the method could also be used to determine both organic and inorganic C content in the soil. A potential disadvantage, however, is that the HCl fumigation changed the ¹⁵N natural abundance of soil N.

Abbreviations: SOM, soil organic matter • FACE, Free Atmospheric CO₂ Enrichment • HCl, HCl acid • ¹³C, natural abundance ¹³C

	INTRODUCTION

TOP ABSTRACT INTRODUCTION Materials and Methods Results and Discussion Conclusions REFERENCES

 
THE ¹³C METHOD is increasingly being used to follow C dynamics in the soil (Balesdent and Mariotti, 1996; Collins et al., 1999). The difference between the ¹³C content of the existing soil organic matter (SOM) and the new plant C may result from changes in the photosynthetic pathway of vegetation (C3 vs. C4) (Balesdent and Mariotti, 1996; Collins et al., 1999), or when ¹³C depleted CO₂ is used to elevate the atmospheric CO₂ concentration such as in a free atmospheric CO₂ enrichment (FACE) experiment (Leavitt et al., 1994; Van Kessel et al., 2000a,b). The difference in the ¹³C values of the new C input and of the older SOM C is sufficiently large to follow the dynamics of both the new and the old C in the soil (Balesdent et al., 1988).

In addition to organic C, soil may also contain inorganic C in the form of carbonates. The C in primary or lithogenic carbonates has ¹³C values close to 0 Pee Dee Belemnite (PDB) (Boutton, 1991). Pedogenic or secondary carbonates show ¹³C values between -12 and +2 (PDB), depending on the photosynthetic pathway of vegetation (C3 vs. C4). When a sample is combusted at high temperature (1000°C), all organic and inorganic C present in the soil is converted into CO₂. To avoid the confounding influence of inorganic C during the determination of the isotopic signature of the organic C, all carbonates must be removed prior to isotopic analysis. Because carbonates may be enriched in ¹³C by as much as 30 compared with organic C, partial removal of carbonates will have a large effect on the ¹³C signature of the sample. For example, if residual carbonate at 0 accounts for 1% of total soil C in a soil where organic C is -25, the inclusion of the CO₂ in the measurement would result in a 0.25 error.

A common method to remove carbonates from soil is treatment with dilute HCl or H₃PO₄ (Connin et al., 1997; Rochette and Flanagan, 1997; Collins et al., 1999; Van Kessel et al., 2000b). Although acid washing will remove all carbonates, the procedure is time consuming and could lead to losses of acid-soluble organic C. Thus, there is a risk that the soluble C may be isotopically different from the insoluble residue. However, Midwood and Boutton (1998) found that the ¹³C signature of SOM C was largely unaffected by the acid concentration (0.1–6 M HCl) or duration (1–8 d) of the acid treatment. To avoid losses of soluble organic C and possible changes in the ¹³C signature of SOM, the removal of carbonates can be carried out by HCl fumigation (Hedges and Stern, 1984).

Using continuous flow isotope ratio mass spectrometers, it has been common to determine both the ¹³C and ¹⁵N isotopes from a single combusted sample (Nadelhoffer and Fry, 1988; Van Kessel et al., 1994). However, the possibility that HCl fumigation changes the ¹⁵N isotopic composition of the sample must be investigated if C and N isotope measurements are to be routinely applied.

The main objectives of this study were (i) to determine the effectiveness of HCl fumigation in removing carbonates from soil, (ii) to test the effects of HCl treatments on the residual SOM ¹³C, and (iii) to determine the effects of the HCl treatment on ¹⁵N of soil N. The possible use of the HCl-fumigation method to determine inorganic and organic C contents was also explored.

	Materials and Methods

TOP ABSTRACT INTRODUCTION Materials and Methods Results and Discussion Conclusions REFERENCES

 
To determine the rate and effectiveness of carbonate removal by HCl vapor, a calcareous soil with a high inorganic C content was selected (Table 1). The calcareous soil (Mollisol) was oven-dried and ball-milled for 24 h. Subsamples (30 mg) of soil were placed in open Ag-foil capsules (8 by 5 mm). Silver capsules are required because Sn capsules disintegrate when exposed to HCl vapor. The capsules were placed in the wells of a microtiter plate, sufficient water was added to each capsule (50 µL) to moisten the soil to approximately field capacity, and the microtiter plate was then placed in a vacuum desiccator (5 L). A beaker (150 mL) with 100 mL of concentrated (12 M) HCl was also placed inside the desiccator. The samples were exposed to HCl vapor for times increasing in 6-h increments to 30 h, then 12-h increments to 96 h. After each exposure time, four replicates were removed from the desiccator, dried at 60°C for 4 h, and the capsules were then closed.

The procedure of Midwood and Boutton (1998) was followed for acid washing the soils. Briefly, 5 g of ball-milled soil was treated with 150 mL of 0.5 M HCl and the soil-acid mixture stirred three times over a 24-h period. Soils were then washed twice with distilled water. Each time the water was replaced after a 24-h period. Soil was dried at 60°C, ground by mortar and pestle, 30-mg samples were weighed into Ag foil capsules, and the capsules closed for isotopic analysis. Four replicates were used for all analyses.

Total N, total C, ¹⁵N, and the ¹³C for all samples were determined on a Europa 20-20 continuous flow isotope ratio mass spectrometer (PDZ Europa Ltd., Sandbach, UK) following combustion at 1000°C in a Europa ANCA-GSL CN analyzer (PDZ Europa Ltd., Sandbach, UK). The ¹⁵N values are expressed relative to atmospheric N₂. The ¹³C values are expressed relative to Vienna-Pee Dee Belemnite (V-PDB). Controls (Ag capsules, HCl fumigation and water) were included. The amounts of C and N in the controls never exceeded 2 µg for N and 6 µg for C, and were too low to obtain a reliable isotopic composition.

One-way analysis of variance followed by the Student-Newman Keuls test (SAS Institute, 1989) was used to test for differences in total N, total C, ¹⁵N, and ¹³C between treatments.

	Results and Discussion

TOP ABSTRACT INTRODUCTION Materials and Methods Results and Discussion Conclusions REFERENCES

 
A common practice for removing carbonates from soil is washing the samples with dilute acid followed by several washings with deionized water (Connin et al., 1997; Midwood and Boutton, 1998; Collins et al., 1999). Although, the ¹³C of the bulk soil organic C may be unaffected by this treatment (Midwood and Boutton, 1998), it is conceivable that the composition of isotopically heterogeneous soils could be altered by this treatment through removal of dissolved organic C. Fumigating the soil with HCl may reduce or eliminate any organic C losses and minimize the potential for changes in ¹³C.

Acid fumigation with HCl of the four noncalcareous soils did not alter their ¹³C compositions or their organic C and total N contents (Table 2). Treating the same soils with dilute HCl followed by washing led to significant declines in total soil C and N for all four soils. For three soils, the ¹³C value also became more negative. As the decline in the ¹³C value for these three soils ranged between 0.09 and 0.20, the removed C was enriched in ¹³C compared with residual organic C. When coralline sediments that contained >80% by weight CaCO₃ were exposed to HCl fumigation, no contamination or loss of organic C and N occurred (Yamamuro and Kayanne, 1995). In contrast, when samples were acidified with 20 mL of 1 M HCl and washed with distilled water, 20% of the C and N was lost. Since those samples were not analyzed for isotopic composition, it remains unknown whether the ¹³C and ¹⁵N abundances were changed. Midwood and Boutton (1998) used one of the carbonate-free soils included in this study (Pophers). When they treated this soil with 1 M HCl for up to 8 d, followed by washing the soil with distilled water, organic C concentration declined significantly but the ¹³C value of the soil was unaffected.

Exposure to concentrated HCl vapor for 6 h completely removed carbonates from a soil that contained high levels of carbonates (Fig. 1) . The high ¹³C value (-5) of untreated soil reflected a strong presence of carbonates. The ¹³C value declined to -27 following HCl fumigation for 6 h. No further decline in the ¹³C value occurred for the remainder of the fumigation period. At the same time, the total C content of the soil declined from a maximum of 32 mg g^-1 to 8 mg g^-1 after 6 h of HCl fumigation and remained constant thereafter (Fig. 1). However, Hedges and Stern (1984) found that HCl-vapor treatment failed to remove all carbonate from a soil with 50% carbonate, suggesting that HCl fumigation may not always be completely effective in removing carbonates from highly calcareous soils.

View larger version (15K): [in this window] [in a new window]   Fig. 1. Changes in soil C concentration and the 13C content of soil C following the removal of carbonates by HCl fumigation. The open squares represent C values, the closed squares represent 13C values.

The HCl fumigation technique is a convenient and easy method to remove inorganic C and could potentially also be used to determine the quantity of organic and inorganic C in a soil. One sample of the soil should be fumigated with HCl, another sample left untreated. Following total C analysis, the difference in total C content in the soil is attributed to carbonates. Total soil C content in the calcareous soil before HCl fumigation was 32 mg g^-1 soil and decreased to 8 mg g^-1 soil after HCl fumigation. Therefore, 24 mg g^-1 soil or 75% of the total C content in the soil was in the form of inorganic C.

It should be pointed out that the HCl-fumigation method for determining inorganic C requires two separate analyses: total organic and inorganic C (untreated sample), and organic C (treated sample). As both analyses contribute experimental error, the calculated amount of inorganic C based on the difference between the C in the two analyses may be less accurate than a direct measurement. However, this point requires further research.

Finally, it should be mentioned that the HCl-fumigation method does reduce, by about 50%, the number of samples that can be combusted before the elemental analyzer combustion reactor is exhausted.

	Conclusions

TOP ABSTRACT INTRODUCTION Materials and Methods Results and Discussion Conclusions REFERENCES

 
Fumigating soil with HCl vapor rather than washing the soil in dilute HCl is an effective method to remove carbonates prior to isotopic analysis. The method does not alter the ¹³C signature of organic C and no losses of organic C occurred. In a soil that contained 75% of total C as carbonates, all carbonates were removed within a 6-h period of fumigation. A large number of samples can be processed simultaneously. However, HCl fumigation may lead to a small increase in the ¹⁵N value of soil N.

	ACKNOWLEDGMENTS

 
We thank Dr. T. Boutton, Texas A&M University, and Dr. M. Singer, University of California-Davis, for providing us with the carbonate-free soil samples. We also thank Chris Hartley for the statistical analyses.

Received for publication July 13, 2000.

	REFERENCES

TOP ABSTRACT INTRODUCTION Materials and Methods Results and Discussion Conclusions REFERENCES

 

• Balesdent, J., and A. Mariotti. 1996. Measurement of soil organic matter turnover using ¹³C natural abundance. p. 83–112. In T.W. Boutton and S. Yamasaki (ed.) Mass spectrometry of soils. Marcel Dekker, New York.
• Balesdent, J., G.H. Wagner, and A. Mariotti. 1988. Soil organic matter turnover in long-term field experiments as revealed by carbon-13 natural abundance. Soil Sci. Soc. Am. J. 52:118–124.[Abstract/Free Full Text]
• Boutton, T.W. 1991. Stable carbon isotope ratios of natural materials: II. Atmospheric, terrestrial, marine, and freshwater environments. p. 173–185. In D.C. Coleman and B. Fry (ed.) Carbon isotope techniques. Academic Press, San Diego, CA.
• Collins, H.P., R.L. Blevins, L.G. Bundy, D.R. Christensen, W.A. Dick, D.R. Huggins, and E.A. Paul. 1999. Soil carbon dynamics in corn-based agroecosystems: Results from carbon-13 natural abundance. Soil Sci. Soc. Am. J. 63:584–591.[Abstract/Free Full Text]
• Connin, S.L., R.A. Virginia, and C.P. Chamberlain. 1997. Carbon isotopes reveal soil organic matter dynamics following arid land shrub expansion. Oecologia 110:374–386.[ISI]
• Hedges, J.L., and J.H. Stern. 1984. Carbon and nitrogen determination of carbonate-containing solids. Limnol. Oceanogr. 29:657–663.
• Leavitt, S.W., E.A. Paul, B.A. Kimball, G.R. Hendrey, J.R. Mauney, R. Rauschkolb, H. Rogers, K.F. Lewin, J. Nagy, P.J. Pinter, Jr., and H.B. Johnson. 1994. Carbon isotope dynamics of free-air CO₂-enriched cotton and soils. Agric. For. Meteor. 70:87–101.
• Lohse, L., R.T. Klooserhuis, H.C. de Stighter, W. Helder, W. van Raaphorst, and T.C.E. van Weering. 2000. Carbonate removal by acidification causes loss of nitrogenous compounds in continental margin sediments. Mar. Chem. 69:193–201.
• Midwood, A.J., and T.W. Boutton. 1998. Soil carbonate decomposition by acid has little effect on the ¹³C or organic matter. Soil Biol. Biochem. 30:1301–1307.
• Munn, J.R., Jr., and M.J. Singer. 1981. Relative erodibility of California benchmark soils. Tech. Rep. USDA–SCS. Land, Air and Water Resources papers 10007, Univ. California-Davis.
• Nadelhoffer, K.J., and B. Fry. 1988. Controls on natural nitrogen-15 and carbon-13 abundance in forest soil organic matter. Soil Sci. Soc. Am. J. 52:1633–1640.[Abstract/Free Full Text]
• Rochette, P., and L.B. Flanagan. 1997. Quantifying rhizosphere respiration in a corn crop under field conditions. Soil Sci. Soc. Am. J. 61:466–474.[Abstract/Free Full Text]
• SAS Institute. 1989. SAS/STAT user's guide. Version 6. 4th ed. Vol. 2. SAS Inst., Cary, NC.
• Trott, K.E. 1981. The importance of selected soil physical and chemical properties for the prediction of soil erodibility of several western upland soils. Ph.D. diss. Dept. of Land, Air and Water Resources, Univ. of California, Davis, CA.
• Van Kessel, C., R.E. Farrell, and D.J. Pennock. 1994. Carbon-13 and nitrogen-15 natural abundance in crop residues and soil organic matter. Soil Sci. Soc. Am. J. 58:382–389.[Abstract/Free Full Text]
• Van Kessel, C., W.R. Horwath, U. Hartwig, D. Harris, and A. Lüscher. 2000a. Net soil carbon input under ambient and elevated CO₂ concentrations: isotopic evidence after four years. Global Change Biol. 6:435–444.
• Van Kessel, C., J. Nitschelm, W.R. Horwath, D. Harris, F. Walley, A. Lüscher, and U. Hartwig. 2000b. Carbon-13 input and turn-over in a pasture soil exposed to long-term elevated atmospheric CO₂. Global Change Biol. 6:123–135.
• Yamamuro, M., and H. Kayanne. 1995. Rapid direct determination of organic carbon and nitrogen in carbonate-bearing sediments with a Yanaco MT-5 CHN analyzer. Limnol. Oceanogr. 40:1001–1005.

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