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Microbial immobilization and mineralization of dissolved organic nitrogen from forest floors
Soil Biology & Biochemistry 43 (2011) 1742e1745
Contents lists available at ScienceDirect
Soil Biology & Biochemistry journal homepage: www.elsevier.com/locate/soilbio
Short Communication
Microbial immobilization and mineralization of dissolved organic nitrogen from forest floors Bettina H.M. Schmidt a, Karsten Kalbitz b, *, Sabine Braun a, Roland Fuß c, d, William H. McDowell e, Egbert Matzner a a
Department of Soil Ecology, Bayreuth Center of Ecology and Environmental Research (BayCEER), University of Bayreuth, 95448 Bayreuth, Germany Earth Surface Science, Institute for Biodiversity and Ecosystem Dynamics, University of Amsterdam, Science Park 904, 1098 XH Amsterdam, The Netherlands Institute of Soil Ecology, Helmholtz Zentrum München, 85764 Neuherberg, Germany d Institute of Agricultural Climate Research, Johann Heinrich von Thünen Institute, 38116 Braunschweig, Germany e Department of Natural Resources and the Environment, University of New Hampshire, Durham, NH 03824, USA b c
a r t i c l e i n f o
a b s t r a c t
Article history: Received 18 March 2011 Received in revised form 21 April 2011 Accepted 27 April 2011 Available online 13 May 2011
Dissolved organic nitrogen (DON) plays a key role in the N cycle of many ecosystems, as DON availability and biodegradation are important for plant growth, microbial metabolism and N transport in soils. However, biodegradation of DON (defined as the sum of mineralization and microbial immobilization) is only poorly understood. In laboratory incubations, biodegradation of DON and dissolved organic carbon (DOC) from Oi and Oa horizons of spruce, beech and cypress forests ranged from 6 to 72%. Biodegradation of DON and DOC was similar in most samples, and mineralization of DON was more important than microbial immobilization. Nitrate additions (0e10 mg N L1) never influenced either DON immobilization by microorganisms or mineralization. We conclude that soil microorganisms do not necessarily prefer mineral N over DON for meeting their N demand, and that biodegradation of DON seems to be driven by the microbial demand for C rather than N. Quantifying the dynamics of DON in soils should include consideration of both C and N demands by microbes. Ó 2011 Elsevier Ltd. All rights reserved.
Keywords: Dissolved organic nitrogen Dissolved organic carbon Nitrate Biodegradation Mineralization Microbial immobilization
Until now, research on the biodegradability of dissolved organic matter (DOM) in soils has focused on the dynamics of dissolved organic carbon (DOC) (e.g. Yano et al., 2000; Marschner and Kalbitz, 2003; Kalbitz et al., 2003; Schwesig et al., 2003; Don and Kalbitz, 2005; Qualls, 2005). Studies concerning the biodegradability of DON in forest floor material or litter leachates are rare (e.g. Qualls and Haines, 1992; Cleveland et al., 2004; Kiikkilä et al., 2005). In addition, it is still uncertain whether DOC and DON behave similarly in soils with respect to biodegradability, although DOC and DON dynamics are strongly correlated in many soils (Ghani et al., 2007). The availability of mineral N might influence DON biodegradation since mineral N may decrease the need of microorganisms to degrade DON. Gregorich et al. (2003) showed that the addition of mineral N (NHþ 4 and NO3 ) to maize-cropped soils did not have any effect on DON biodegradation in water extracts of these soils. Yano
* Corresponding author. Tel.: þ31 20 525 7457; fax: þ31 20 525 7432. E-mail addresses: [email protected] (B.H.M. Schmidt), [email protected] (K. Kalbitz), [email protected] (S. Braun), [email protected] (R. Fuß), bill. [email protected] (W.H. McDowell), [email protected] (E. Matzner). 0038-0717/$ e see front matter Ó 2011 Elsevier Ltd. All rights reserved. doi:10.1016/j.soilbio.2011.04.021
et al. (2000) found an increase in biodegradable DOC concentrations in field-derived solutions after NH4NO3 additions to a hardwood stand. In contrast, there is no study that deals with the influence of mineral N availability on DON biodegradation in forest floors. In this study, biodegradation of DON is referred to as the sum of mineralization (use of organic compounds as a source of energy and nutrients) and microbial immobilization (formation of particulate organic matter, POM). Thus, DON (and DOC) biodegradation can be calculated as the difference between initial and final DON (and DOC) concentrations. Microbial immobilization of DON (e.g. amino acids) results in an increase of particulate organic nitrogen (PON). If DON in solution is decomposed by microbes, NHþ 4 concentrations will increase (mineralization). In turn, NHþ 4 can be oxidized to NO3 by nitrification, but NHþ can also be incorporated into microbial 4 biomass (increase in PON) and DON can be released by (dead) microbial biomass (decrease in PON). We hypothesized that (i) DOC and DON biodegradation are similar and that (ii) NO 3 availability is an important control of DON biodegradation and increasing its supply would lead to a decrease in DON biodegradation, particularly in samples with high C availability (Geisseler et al., 2010).
B.H.M. Schmidt et al. / Soil Biology & Biochemistry 43 (2011) 1742e1745
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Table 1 þ Chemical characteristics (pH, DOC and DON concentration, DOC/DON ratio, NO 3 and NH4 concentrations and specific UV absorbance at 280 nm - SUVA280) of the five different water extracts from Oi and Oa horizons of beech, spruce and cypress (n.d.: not detectable). Numbers in brackets are standard deviations (n ¼ 3).
Beech-Oi Beech-Oa Spruce-Oi Spruce-Oa Cypress-Oa
pH
DON [mg N L1]
DOC [mg C L1]
DOC/DON
NO 3 [mg N L1]
NHþ 4 [mg N L1]
SUVA (280 nm) [L 10 mg C1 cm1]
6.3 5.8 5.4 4.3 4.7
2.00 3.21 4.47 1.08 0.57
71.18 78.53 81.47 24.48 11.80
35.56 24.46 18.24 22.60 20.88
n. d. 0.31 (0.04) n. d. n. d. n. d.
0.74 (0.14) 4.27 (0.06) 3.28 (0.03 0.60 (0.01) 1.06 (0.03)
0.26 (0.01) 0.28 (0.01) 0.09 (0.00) 0.33 (0.01) 0.26 (0.03)
(0.04) (0.04) (0.18) (0.03) (0.02)
(0.94) (0.66) (0.91) (0.75) (0.22)
Soil samples were collected from the Oi and Oa horizons of three long-term ecological research sites with a different N status and availability as revealed by DOC/DON ratios in throughfall and forest floor percolates and fluxes of mineral and organic N in the respective ecosystems (Schmidt and Matzner, 2009): the Chi-Lan Mountain forest ecosystem in Northern Taiwan (cypress, Chamaecyparis formosana var. obtusa, only Oa material, highest DOC/DON ratios and lowest mineral N fluxes in throughfall), the Steinkreuz site (European beech, Fagus sylvatica L.) and the Coulissenhieb site (Norway spruce, Picea abies (L.) Karst., highest mineral N fluxes in throughfall), both in Germany. Details concerning climate, soils and vegetation can be found in Gerstberger et al. (2004) and Schmidt et al. (2010); the chemical characteristics of the extracts are listed in Table 1.
6
80
4
60
3
40
2 20
0
0 0
-1
80
4
60
3
40
5
10
15
0
20 6
spruce-Oi
80
5 4
60
3
40
20
1
0 0
5
10
15
20
spruce-Oa
80
5 4
60
3
40
2
2 1 0 0
5
10
15
20
1
0
0
cypress-Oa
4 3
0 0
20
5
20
5
80
DON + NH4
60
PON DOC
10
15
DOC concentration [mg C L-1]
N concentration [mg N L ]
beech-Oa
5
2
1
6
Dissolved organic matter was prepared according to Kalbitz et al. (2003) by adding 3 L of water to 300 g fresh weight of litter horizon material. The suspensions were stirred three times during the extraction period (24 h, 5 C). Solutions were filtered through 0.45 mm prewashed cellulose acetate filters (Whatman, OE67). Samples of all Oa horizons were used to prepare a mixed inoculum, which was added to each DOM solution (ratio 1:100, for details see Kalbitz et al., 2003). Mineral N was added as NaNO3 (0, 3, 1 for beech-Oi, and 0, 0.75, 1.5 and 3 mg 5 and 10 mg NO 3 eN L 1 NO3 eN L for all other solutions) to obtain similar DOC/TDN (total dissolved N) ratios. Controls with pure water instead of DOM solutions were treated in the same way to assess C and N losses from the inoculum. All samples were incubated in triplicates in closed PE bottles at 20 C in the dark for 21 days. Subsamples were
6
beech-Oi
5
6
(0.95) (0.10) (0.61) (0.57) (0.40)
20
40
2 20
1 0
0 0
5
10
15
20
incubation time [days] Fig. 1. Changes in DOC, DON, and PON concentrations in water extracts of beech-Oi, beech-Oa, spruce-Oi, spruce-Oa and cypress-Oa during 21-day incubations without NO 3 addition. Error bars represent one standard deviation (n ¼ 3). NHþ 4
1744
B.H.M. Schmidt et al. / Soil Biology & Biochemistry 43 (2011) 1742e1745
5 beech-Oi*
beech-Oa
4
4
3
3
2
2
1
1
0
0
0
5
10
15
20
5 spruce-Oi
-1
DON concentration [mg N L ]
5
0
10
15
20
5 spruce-Oa
4
4
3
3
2
2
1
1
0
5
0 0
5
10
15
20
0
5
10
15
20
5 cypress-Oa 4 3
-1
0 / 0* mg N L -1 0.75 / 3* mg N L -1 1.5 / 5* mg N L -1 3 / 10* mg N L
2 1 0 0
5
10
15
20
incubation time [days] Fig. 2. Changes in DON concentrations in water extracts of beech-Oi, beech-Oa, spruce-Oi, spruce-Oa and cypress-Oa during 21-day incubations with NO 3 addition. NO3 additions with asterisks (*) are the rates applied to the beech-Oi-samples only. Error bars represent one standard deviation (n ¼ 3).
taken after 0, 1, 3, 5, 7, 10, 14 and 21 days. Before sampling, solutions were gently shaken to homogenize and aerate the samples. In filtered subsamples (Minisart, 0.45 mm, cellulose acetate, prewashed), TDN, DOC (multi N/C 2100, Analytik Jena, detection limit TDN: 0.15 mg N L1, NPOC: 0.66 mg C L1), NO 3 (Dionex DX 500 ion chromatograph, detection limit 0.29 mg N L1) and NHþ 4 (FIA-LAB flow injector, MLE Dresden, detection limit: 0.25 mg N L1) were determined. In addition, TN was also determined on unfiltered subsamples. Dissolved organic N was calculated from filtered subsamples as TDNðNHþ 4 þ NO3 Þ. Nitrite was assumed to be negligible. Particulate organic N was calculated as the difference in TN between filtered and unfiltered samples and used as proxy for microbial biomass N. Specific UV absorbance (SUVA280) was measured with a UV spectrophotometer at 280 nm (UV-1800, Shimadzu). The temporal dynamics of DOC and DON biodegradation were similar (Fig. 1). Dissolved organic N biodegradation was higher than DOC biodegradation in spruce-Oi and beech-Oa-samples, while
DOC and DON biodegradation were similar in all other samples. Dissolved organic C and N biodegradation ranged from 6 to 63% and 6 to 72%, respectively, and were highest in extracts from spruce-Oihorizons, while in spruce-Oa and cypress-Oa-samples, DOC and DON concentrations were stable. Nitrogen mineralization (i.e. increase in NHþ 4 ) was always larger than microbial N immobilization (i.e. increase in PON). The increase in PON was negligible in Oasamples and similar in both Oi-samples. Previous work has shown that DOC and DON can follow both similar (Cleveland et al., 2004; Kiikkilä et al., 2005; Qualls and Haines, 1992) and divergent trajectories during biodegradation of DOM (Gregorich et al., 2003). This divergence is possible because dissolved organic C and N may be concentrated in different fractions of DOM (hydrophobic and hydrophilic, respectively) (Kaiser and Zech, 2000; Petrone et al., 2009) and differential utilization of these fractions might cause divergence in DOC and DON biodegradation. We found particularly large DOC and DON biodegradation in samples with low specific UV absorbance,
B.H.M. Schmidt et al. / Soil Biology & Biochemistry 43 (2011) 1742e1745
indicating a largely hydrophilic character of the organic matter in these samples. Generally, hydrophilic components are more rapidly degraded than hydrophobic ones (Kalbitz et al., 2003). Qualls and Haines (1992) postulated that the hydrolysis of DON is closely linked to DOC mineralization, and is not driven by a biochemical need of microorganisms for N. NO 3 additions, regardless of magnitude, had no effect on DON concentrations or dynamics in any of the solutions (Fig. 2). The same was true for DOC and PON (data not shown). NHþ 4 concentrations only increased at the highest NO 3 addition in Oi-samples (data not shown). We had hypothesized that addition of mineral N would decrease the biodegradation of DON due to a preferential uptake of mineral N forms. This hypothesis was not supported by our data, as DON biodegradation never decreased after NO 3 addition, although overall rates of DOC and DON biodegradation in our samples covered a wide range. Thus, we conclude that DON and DOC biodegradation were not limited by N. We did observe that the highest DON biodegradation occurred in samples with high initial NHþ 4 concentrations (spruce-Oi, Table 1). Because the influence of NHþ 4 concentration on DON biodegradation was not tested systematically in this study, we are not certain if there is a strong relationship between NHþ 4 concentrations and DON biodegradation. Microorganisms in our incubations were unlikely to be limited by P or other nutrients, as analyses in water extracts from our sites concentrations, for example, between 2.3 and yielded PO3 4 9.7 mg P L1. Although characterization of the DOC and DON in organic matter is useful for assessment of C and N fluxes in forest ecosystems, our results suggest that DOC and DON should be considered as a single pool of dissolved organic matter when undergoing biodegradation, as DOC and DON biodegradation showed similar temporal dynamics in all samples. Dissolved organic C and N are two parameters that can be used to quantify and characterize DOM, such as through the C/N ratio, but they do not represent distinct compounds that behave differently in forest ecosystems. Despite the fact that our samples varied in initial chemical properties and came from forests having a different N status and availability, addition of inorganic N did not affect DON biodegradation. This suggests that DON biodegradation is driven by the C demand of microorganisms, rather than N availability. Acknowledgments This study was funded by the Deutsche Forschungsgemeinschaft, Germany. The authors would like to thank Prof. Dr. Shih-Chieh Chang for providing samples from the Chi-Lan Mountain site,
1745
Taiwan and Gerard Ros for helpful comments on an earlier version of this manuscript. We are grateful to the members of the Central Analytical Department of the Bayreuth Center of Ecology and Environmental Research (BayCEER) for their help with the analysis of samples.
References Cleveland, C.C., Neff, J.C., Townsend, A.R., Hood, E., 2004. Composition, dynamics, and fate of leached dissolved organic matter in terrestrial ecosystems: results from a decomposition experiment. Ecosystems 7, 275e285. Don, A., Kalbitz, K., 2005. Amounts and degradability of dissolved organic carbon from foliar litter at different decomposition stages. Soil Biology & Biochemistry 37, 2171e2179. Geisseler, D., Horwartg, W.R., Jörgensen, R., Ludwig, B., 2010. Pathways of nitrogen utilization by soil microorganisms - A review. Soil Biology & Biochemistry 42, 2058e2067. Gerstberger, P., Foken, T., Kalbitz, K., 2004. The Lehstenbach and Steinkreuz Catchments in NE Bavaria, Germany. In: Matzner, E. (Ed.), Biogeochemistry of Forested Catchments in a Changing Environment. A German Case Study. Springer, Berlin, pp. 15e41. Ghani, A., Dexter, M., Carran, R.A., Theobald, P.W., 2007. Dissolved organic nitrogen and carbon in pastoral soils: the New Zealand experience. European Journal of Soil Science 58, 832e843. Gregorich, E.G., Beare, M.H., Stoklas, U., St-Georges, P., 2003. Biodegradability of soluble organic matter in maize-cropped soils. Geoderma 113, 237e252. Kaiser, K., Zech, W., 2000. Sorption of dissolved organic nitrogen by acid subsoil horizons and individual mineral phases. European Journal of Soil Science 51, 403e411. Kalbitz, K., Schmerwitz, J., Schwesig, D., Matzner, E., 2003. Biodegradation of soilderived dissolved organic matter as related to its properties. Geoderma 113, 273e291. Kiikkilä, O., Kitunen, V., Smolander, A., 2005. Degradability of dissolved soil organic carbon and nitrogen in relation to tree species. FEMS Microbiology Ecology 53, 33e40. Marschner, B., Kalbitz, K., 2003. Controls of bioavailability and biodegradability of dissolved organic matter in soils. Geoderma 113, 211e235. Petrone, K.C., Richards, J.S., Grierson, P.F., 2009. Bioavailability and composition of dissolved organic carbon and nitrogen in a near coastal catchment of southwestern Australia. Biogeochemistry 92, 27e40. Qualls, R.G., 2005. Biodegradability of fractions of dissolved organic carbon leached from decomposing leaf litter. Environmental Science & Technology 39, 1616e1622. Qualls, R.G., Haines, B.L., 1992. Biodegradability of dissolved organic matter in forest throughfall, soil solution, and stream water. Soil Science Society of America Journal 56, 578e586. Schmidt, B.H.M., Matzner, E., 2009. Abiotic reaction of nitrite with dissolved organic carbon? Testing the Ferrous Wheel Hypothesis. Biogeochemistry 93, 291e296. Schmidt, B.H.M., Wang, C.-P., Chang, S.-C., Matzner, E., 2010. High precipitation causes large fluxes of dissolved organic carbon and nitrogen in a subtropical montane Chamaecyparis forest in Taiwan. Biogeochemistry 101, 243e256. Schwesig, D., Kalbitz, K., Matzner, E., 2003. Mineralization of dissolved organic carbon in mineral soil solution of two forest soils. Journal of Plant Nutrition and Soil Science 166, 585e593. Yano, Y., McDowell, W.H., Aber, J.D., 2000. Biodegradable dissolved organic carbon in forest soil solution and effects of chronic nitrogen deposition. Soil Biology & Biochemistry 32, 1743e1751.
Contents lists available at ScienceDirect
Soil Biology & Biochemistry journal homepage: www.elsevier.com/locate/soilbio
Short Communication
Microbial immobilization and mineralization of dissolved organic nitrogen from forest floors Bettina H.M. Schmidt a, Karsten Kalbitz b, *, Sabine Braun a, Roland Fuß c, d, William H. McDowell e, Egbert Matzner a a
Department of Soil Ecology, Bayreuth Center of Ecology and Environmental Research (BayCEER), University of Bayreuth, 95448 Bayreuth, Germany Earth Surface Science, Institute for Biodiversity and Ecosystem Dynamics, University of Amsterdam, Science Park 904, 1098 XH Amsterdam, The Netherlands Institute of Soil Ecology, Helmholtz Zentrum München, 85764 Neuherberg, Germany d Institute of Agricultural Climate Research, Johann Heinrich von Thünen Institute, 38116 Braunschweig, Germany e Department of Natural Resources and the Environment, University of New Hampshire, Durham, NH 03824, USA b c
a r t i c l e i n f o
a b s t r a c t
Article history: Received 18 March 2011 Received in revised form 21 April 2011 Accepted 27 April 2011 Available online 13 May 2011
Dissolved organic nitrogen (DON) plays a key role in the N cycle of many ecosystems, as DON availability and biodegradation are important for plant growth, microbial metabolism and N transport in soils. However, biodegradation of DON (defined as the sum of mineralization and microbial immobilization) is only poorly understood. In laboratory incubations, biodegradation of DON and dissolved organic carbon (DOC) from Oi and Oa horizons of spruce, beech and cypress forests ranged from 6 to 72%. Biodegradation of DON and DOC was similar in most samples, and mineralization of DON was more important than microbial immobilization. Nitrate additions (0e10 mg N L1) never influenced either DON immobilization by microorganisms or mineralization. We conclude that soil microorganisms do not necessarily prefer mineral N over DON for meeting their N demand, and that biodegradation of DON seems to be driven by the microbial demand for C rather than N. Quantifying the dynamics of DON in soils should include consideration of both C and N demands by microbes. Ó 2011 Elsevier Ltd. All rights reserved.
Keywords: Dissolved organic nitrogen Dissolved organic carbon Nitrate Biodegradation Mineralization Microbial immobilization
Until now, research on the biodegradability of dissolved organic matter (DOM) in soils has focused on the dynamics of dissolved organic carbon (DOC) (e.g. Yano et al., 2000; Marschner and Kalbitz, 2003; Kalbitz et al., 2003; Schwesig et al., 2003; Don and Kalbitz, 2005; Qualls, 2005). Studies concerning the biodegradability of DON in forest floor material or litter leachates are rare (e.g. Qualls and Haines, 1992; Cleveland et al., 2004; Kiikkilä et al., 2005). In addition, it is still uncertain whether DOC and DON behave similarly in soils with respect to biodegradability, although DOC and DON dynamics are strongly correlated in many soils (Ghani et al., 2007). The availability of mineral N might influence DON biodegradation since mineral N may decrease the need of microorganisms to degrade DON. Gregorich et al. (2003) showed that the addition of mineral N (NHþ 4 and NO3 ) to maize-cropped soils did not have any effect on DON biodegradation in water extracts of these soils. Yano
* Corresponding author. Tel.: þ31 20 525 7457; fax: þ31 20 525 7432. E-mail addresses: [email protected] (B.H.M. Schmidt), [email protected] (K. Kalbitz), [email protected] (S. Braun), [email protected] (R. Fuß), bill. [email protected] (W.H. McDowell), [email protected] (E. Matzner). 0038-0717/$ e see front matter Ó 2011 Elsevier Ltd. All rights reserved. doi:10.1016/j.soilbio.2011.04.021
et al. (2000) found an increase in biodegradable DOC concentrations in field-derived solutions after NH4NO3 additions to a hardwood stand. In contrast, there is no study that deals with the influence of mineral N availability on DON biodegradation in forest floors. In this study, biodegradation of DON is referred to as the sum of mineralization (use of organic compounds as a source of energy and nutrients) and microbial immobilization (formation of particulate organic matter, POM). Thus, DON (and DOC) biodegradation can be calculated as the difference between initial and final DON (and DOC) concentrations. Microbial immobilization of DON (e.g. amino acids) results in an increase of particulate organic nitrogen (PON). If DON in solution is decomposed by microbes, NHþ 4 concentrations will increase (mineralization). In turn, NHþ 4 can be oxidized to NO3 by nitrification, but NHþ can also be incorporated into microbial 4 biomass (increase in PON) and DON can be released by (dead) microbial biomass (decrease in PON). We hypothesized that (i) DOC and DON biodegradation are similar and that (ii) NO 3 availability is an important control of DON biodegradation and increasing its supply would lead to a decrease in DON biodegradation, particularly in samples with high C availability (Geisseler et al., 2010).
B.H.M. Schmidt et al. / Soil Biology & Biochemistry 43 (2011) 1742e1745
1743
Table 1 þ Chemical characteristics (pH, DOC and DON concentration, DOC/DON ratio, NO 3 and NH4 concentrations and specific UV absorbance at 280 nm - SUVA280) of the five different water extracts from Oi and Oa horizons of beech, spruce and cypress (n.d.: not detectable). Numbers in brackets are standard deviations (n ¼ 3).
Beech-Oi Beech-Oa Spruce-Oi Spruce-Oa Cypress-Oa
pH
DON [mg N L1]
DOC [mg C L1]
DOC/DON
NO 3 [mg N L1]
NHþ 4 [mg N L1]
SUVA (280 nm) [L 10 mg C1 cm1]
6.3 5.8 5.4 4.3 4.7
2.00 3.21 4.47 1.08 0.57
71.18 78.53 81.47 24.48 11.80
35.56 24.46 18.24 22.60 20.88
n. d. 0.31 (0.04) n. d. n. d. n. d.
0.74 (0.14) 4.27 (0.06) 3.28 (0.03 0.60 (0.01) 1.06 (0.03)
0.26 (0.01) 0.28 (0.01) 0.09 (0.00) 0.33 (0.01) 0.26 (0.03)
(0.04) (0.04) (0.18) (0.03) (0.02)
(0.94) (0.66) (0.91) (0.75) (0.22)
Soil samples were collected from the Oi and Oa horizons of three long-term ecological research sites with a different N status and availability as revealed by DOC/DON ratios in throughfall and forest floor percolates and fluxes of mineral and organic N in the respective ecosystems (Schmidt and Matzner, 2009): the Chi-Lan Mountain forest ecosystem in Northern Taiwan (cypress, Chamaecyparis formosana var. obtusa, only Oa material, highest DOC/DON ratios and lowest mineral N fluxes in throughfall), the Steinkreuz site (European beech, Fagus sylvatica L.) and the Coulissenhieb site (Norway spruce, Picea abies (L.) Karst., highest mineral N fluxes in throughfall), both in Germany. Details concerning climate, soils and vegetation can be found in Gerstberger et al. (2004) and Schmidt et al. (2010); the chemical characteristics of the extracts are listed in Table 1.
6
80
4
60
3
40
2 20
0
0 0
-1
80
4
60
3
40
5
10
15
0
20 6
spruce-Oi
80
5 4
60
3
40
20
1
0 0
5
10
15
20
spruce-Oa
80
5 4
60
3
40
2
2 1 0 0
5
10
15
20
1
0
0
cypress-Oa
4 3
0 0
20
5
20
5
80
DON + NH4
60
PON DOC
10
15
DOC concentration [mg C L-1]
N concentration [mg N L ]
beech-Oa
5
2
1
6
Dissolved organic matter was prepared according to Kalbitz et al. (2003) by adding 3 L of water to 300 g fresh weight of litter horizon material. The suspensions were stirred three times during the extraction period (24 h, 5 C). Solutions were filtered through 0.45 mm prewashed cellulose acetate filters (Whatman, OE67). Samples of all Oa horizons were used to prepare a mixed inoculum, which was added to each DOM solution (ratio 1:100, for details see Kalbitz et al., 2003). Mineral N was added as NaNO3 (0, 3, 1 for beech-Oi, and 0, 0.75, 1.5 and 3 mg 5 and 10 mg NO 3 eN L 1 NO3 eN L for all other solutions) to obtain similar DOC/TDN (total dissolved N) ratios. Controls with pure water instead of DOM solutions were treated in the same way to assess C and N losses from the inoculum. All samples were incubated in triplicates in closed PE bottles at 20 C in the dark for 21 days. Subsamples were
6
beech-Oi
5
6
(0.95) (0.10) (0.61) (0.57) (0.40)
20
40
2 20
1 0
0 0
5
10
15
20
incubation time [days] Fig. 1. Changes in DOC, DON, and PON concentrations in water extracts of beech-Oi, beech-Oa, spruce-Oi, spruce-Oa and cypress-Oa during 21-day incubations without NO 3 addition. Error bars represent one standard deviation (n ¼ 3). NHþ 4
1744
B.H.M. Schmidt et al. / Soil Biology & Biochemistry 43 (2011) 1742e1745
5 beech-Oi*
beech-Oa
4
4
3
3
2
2
1
1
0
0
0
5
10
15
20
5 spruce-Oi
-1
DON concentration [mg N L ]
5
0
10
15
20
5 spruce-Oa
4
4
3
3
2
2
1
1
0
5
0 0
5
10
15
20
0
5
10
15
20
5 cypress-Oa 4 3
-1
0 / 0* mg N L -1 0.75 / 3* mg N L -1 1.5 / 5* mg N L -1 3 / 10* mg N L
2 1 0 0
5
10
15
20
incubation time [days] Fig. 2. Changes in DON concentrations in water extracts of beech-Oi, beech-Oa, spruce-Oi, spruce-Oa and cypress-Oa during 21-day incubations with NO 3 addition. NO3 additions with asterisks (*) are the rates applied to the beech-Oi-samples only. Error bars represent one standard deviation (n ¼ 3).
taken after 0, 1, 3, 5, 7, 10, 14 and 21 days. Before sampling, solutions were gently shaken to homogenize and aerate the samples. In filtered subsamples (Minisart, 0.45 mm, cellulose acetate, prewashed), TDN, DOC (multi N/C 2100, Analytik Jena, detection limit TDN: 0.15 mg N L1, NPOC: 0.66 mg C L1), NO 3 (Dionex DX 500 ion chromatograph, detection limit 0.29 mg N L1) and NHþ 4 (FIA-LAB flow injector, MLE Dresden, detection limit: 0.25 mg N L1) were determined. In addition, TN was also determined on unfiltered subsamples. Dissolved organic N was calculated from filtered subsamples as TDNðNHþ 4 þ NO3 Þ. Nitrite was assumed to be negligible. Particulate organic N was calculated as the difference in TN between filtered and unfiltered samples and used as proxy for microbial biomass N. Specific UV absorbance (SUVA280) was measured with a UV spectrophotometer at 280 nm (UV-1800, Shimadzu). The temporal dynamics of DOC and DON biodegradation were similar (Fig. 1). Dissolved organic N biodegradation was higher than DOC biodegradation in spruce-Oi and beech-Oa-samples, while
DOC and DON biodegradation were similar in all other samples. Dissolved organic C and N biodegradation ranged from 6 to 63% and 6 to 72%, respectively, and were highest in extracts from spruce-Oihorizons, while in spruce-Oa and cypress-Oa-samples, DOC and DON concentrations were stable. Nitrogen mineralization (i.e. increase in NHþ 4 ) was always larger than microbial N immobilization (i.e. increase in PON). The increase in PON was negligible in Oasamples and similar in both Oi-samples. Previous work has shown that DOC and DON can follow both similar (Cleveland et al., 2004; Kiikkilä et al., 2005; Qualls and Haines, 1992) and divergent trajectories during biodegradation of DOM (Gregorich et al., 2003). This divergence is possible because dissolved organic C and N may be concentrated in different fractions of DOM (hydrophobic and hydrophilic, respectively) (Kaiser and Zech, 2000; Petrone et al., 2009) and differential utilization of these fractions might cause divergence in DOC and DON biodegradation. We found particularly large DOC and DON biodegradation in samples with low specific UV absorbance,
B.H.M. Schmidt et al. / Soil Biology & Biochemistry 43 (2011) 1742e1745
indicating a largely hydrophilic character of the organic matter in these samples. Generally, hydrophilic components are more rapidly degraded than hydrophobic ones (Kalbitz et al., 2003). Qualls and Haines (1992) postulated that the hydrolysis of DON is closely linked to DOC mineralization, and is not driven by a biochemical need of microorganisms for N. NO 3 additions, regardless of magnitude, had no effect on DON concentrations or dynamics in any of the solutions (Fig. 2). The same was true for DOC and PON (data not shown). NHþ 4 concentrations only increased at the highest NO 3 addition in Oi-samples (data not shown). We had hypothesized that addition of mineral N would decrease the biodegradation of DON due to a preferential uptake of mineral N forms. This hypothesis was not supported by our data, as DON biodegradation never decreased after NO 3 addition, although overall rates of DOC and DON biodegradation in our samples covered a wide range. Thus, we conclude that DON and DOC biodegradation were not limited by N. We did observe that the highest DON biodegradation occurred in samples with high initial NHþ 4 concentrations (spruce-Oi, Table 1). Because the influence of NHþ 4 concentration on DON biodegradation was not tested systematically in this study, we are not certain if there is a strong relationship between NHþ 4 concentrations and DON biodegradation. Microorganisms in our incubations were unlikely to be limited by P or other nutrients, as analyses in water extracts from our sites concentrations, for example, between 2.3 and yielded PO3 4 9.7 mg P L1. Although characterization of the DOC and DON in organic matter is useful for assessment of C and N fluxes in forest ecosystems, our results suggest that DOC and DON should be considered as a single pool of dissolved organic matter when undergoing biodegradation, as DOC and DON biodegradation showed similar temporal dynamics in all samples. Dissolved organic C and N are two parameters that can be used to quantify and characterize DOM, such as through the C/N ratio, but they do not represent distinct compounds that behave differently in forest ecosystems. Despite the fact that our samples varied in initial chemical properties and came from forests having a different N status and availability, addition of inorganic N did not affect DON biodegradation. This suggests that DON biodegradation is driven by the C demand of microorganisms, rather than N availability. Acknowledgments This study was funded by the Deutsche Forschungsgemeinschaft, Germany. The authors would like to thank Prof. Dr. Shih-Chieh Chang for providing samples from the Chi-Lan Mountain site,
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Taiwan and Gerard Ros for helpful comments on an earlier version of this manuscript. We are grateful to the members of the Central Analytical Department of the Bayreuth Center of Ecology and Environmental Research (BayCEER) for their help with the analysis of samples.
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