Bjarne W. Strobel

Influence of vegetation on low-molecular-weight carboxylic acids in soil solution: a review

Low-molecular-weight (LMW) carboxylic acids found in soils and soil solutions comprise mainly aliphatic mono-, di- and tricarboxylic acids and substituted benzoic and cinnamic acids. This review compiles current information on the content of LMW carboxylic acids in soil solutions collected by centrifugation and in lysimeters, and soil extracts in relation to type of vegetation, soil type and soil depth. Contents of LMW carboxylic acids are highest in soil solution from the upper soil layers where carbon in LMW carboxylic acids constitutes up to 10% of DOC. The concentrations of aliphatic LMW di-/tricarboxylic acids (oxalic, malonic, malic, succinic, tartaric and citric acid) are usually in the range 0–50 μM with the higher concentrations in soil solutions from meadow, permanent pasture and forest soils as compared to ley and cultivated soils. In contrast, the concentrations of aliphatic LMW monocarboxylic acids (formic, acetic, propionic, butyric, valeric and lactic acid) are commonly in the range 0–1 mM with high concentrations found in soil solutions from both cultivated and forested soils. Especially insence cedar (Calocedrus decurrens (Torr.) Florin), ponderosa pine (Pinus ponderosa Dougl.), Douglas fir (Pseudotsuga menziesii Franco) and Norway spruce (Picea abies (L.) Karst.) seem to cause higher contents of LMW carboxylic acids in soils and soil solutions as compared to other tree species. Soil solution concentrations of substituted benzoic and cinnamic acids are submicromolar for all vegetations, whereas similar amounts in the range 0–800 μmol kg−1 of aliphatic LMW carboxylic acids and substituted benzoic and cinnamic acids are extractable from mineral soils with alkaline and dilute acid extractants. Seasonal variations are observed in soil solutions isolated by centrifugation and soil extracts with highest concentrations in 2–4 months in a period from mid spring to early summer, minimum concentrations around early autumn, and increasing concentrations during autumn.

Composition and reactivity of DOC in forest floor soil solutions in relation to tree species and soil type

Metal coordinating propertiesof DOC (dissolved organic carbon), and henceits influence on heavy metal release andmineral weathering, depend on the compositionand properties of DOC. Tree species producelitter with different chemical composition anddegradability, and these differences mightinfluence the composition and reactivity of DOCin soil solutions. Accordingly, analysis ofcomposition and reactivity of DOC in soilsolution samples collected by centrifugationfrom 16 forest soil O horizons from the fourtree species beech (Fagus sylvatica L.),oak (Quercus robur L.), grand fir (Abies grandis Lindl.), and Norway spruce (Picea abies (L.) Karst.) on two clayey and twosandy soils were carried out. The compositionand properties of DOC were determined bycapillary zone electrophoresis, acid-basetitration, Cu ion titration, total elementalanalysis, IR and UV spectroscopy, and metalrelease assays. Concentrations of DOC rangedfrom 20 to 163 mM with pH ranging from 4.6 to7.3. Norway spruce produced the highest DOCconcentration, and the lowest pH. Carbon inlow-molecular-weight aliphatic carboxylic acids(LACA) accounted for less than 6% of DOC withformic and acetic acids as the most abundantLACAs. The DOC was cation exchanged and protonsaturated to obtain comparable forms of DOC.Titratable carboxylic acid and phenolic groupswere in the range 51 to 82 and 20 to64 mmol·mol−1 C, respectively, with fewerphenolic groups in grand fir DOC as the onlysignificant difference. Infrared spectra offreeze-dried DOC samples suggest low contentsof aromatic C in the DOC especially from grandfir stands. Stability constants, log K of Cu-DOC complexes, determined by Cu ion titrationof DOC samples with fitting of the data to atwo-site binding model, were in the range 5.63to 6.21 for the strong binding sites and 3.58to 4.10 for the weak sites, but with nosignificant effects of tree species or site.Freeze-dried DOC samples were found to consistof 41 to 45% C, 38 to 49% O, 4.4 to 5.4% Hand 1.2 to 2.0% N and C/N ratios in the range26 to 42. Reactivity of DOC in terms of releaserates of Cd, Cu and Fe cations from a soilsample (flow cell experiments) showed nosignificant differences among DOC samples fromdifferent tree species and soil types.Apparently, only minor differences occur inchemical composition and reactivity ofequivalent concentrations of DOC in forestfloor soil solutions irrespective of origin,i.e. four tree species and two soil types. Soilsolution pH and the concentration of DOCproduced by various tree species are thecritical parameters when distinguishing amongtree species in relation to heavy metal releaseand mineral weathering.

Direct analysis of tfdA gene expression by indigenous bacteria in phenoxy acid amended agricultural soil.

Expression of the functional gene tfdA involved in degradation of phenoxyacetic acids such as 2,4-dichlorophenoxyacetic acid (2,4-D) and 4-chloro-2-methylphenoxyacetic acid (MCPA) was investigated during degradation scenarios in natural unseeded soil samples. The results illustrate how messenger RNA (mRNA)-based analysis is well suited to quantitatively study the activity of specific microbial populations in soil using phenoxyacetic acid biodegradation as a model system. Via quantitative real-time PCR, a clear response to the presence of phenoxy acids was shown during degradation in soil amended with 20 mg 2,4-D or MCPA per kg soil. Further, we found a relatively high degree of correlation between expression of the functional gene and the rates of mineralization. Melting curve analyses of real-time PCR products, supported by tfdA-denaturing gradient gel electrophoresis analysis showed that, although only class I tfdA genes were apparent in the indigenous microbial population, class III tfdA genes became predominant during incubation, and were the only genes expressed during degradation of MCPA in soil. In contrast, both classes were expressed during degradation of the structurally similar compound 2,4-D. The ability to quantify microbial transcripts directly in environmental samples will have a profound impact on our understanding of microbial processes in the environment in future studies.

Cadmium and copper release kinetics in relation to afforestation of cultivated soil

Afforestation of cultivated soils causes soil acidification and elevated concentrations of dissolved organic matter (DOC) in the soil solution, and hence, aggravate the risk of heavy metal leaching. The kinetics of cadmium and copper release from an unpolluted arable soil applied with forest floor soil solution was investigated in the laboratory, and the release rates correlated to pH and DOC in solution through log-log equations. The soil solution was isolated from Norway spruce (Picea abies (L.) Karst.) by centrifugation, and the solution passed a cation-exchange column to remove metal cations and to protonate the DOC. Soil samples from an arable Ap horizon were placed in completely mixed flow cells, and influent solutions with 0 to 5 mM DOC were applied. The solution pH was adjusted to achieve effluent pH values in the range 3.6 to 6.9 in the flow cells at steady-state conditions. Cadmium release rates were very low at pH . 5 and increased exponentially as pH decreased to ,5. The release rate was correlated to solution pH in a simple model: log(cadmium release rate) 52 0.21 pH 2 15.28 (R 2 5 0.48), and no significant effect of DOC was observed. The kinetics of copper release from the soil was more complicated with effects of both pH and DOC. In experiments without DOC, the release rate of copper was slightly lower at high pH than at low pH. In experiments above pH 5, the presence of 5 mM DOC in the solution increased the release rate of copper. However, the copper release was retarded by DOC in the range pH 3.8 to 5.0, which coincided with a maximum retention of DOC in the flow cells. The release rate of copper was correlated to solution pH and concentration of DOC, including an interaction of pH and DOC: log(copper release rate) 5 0.86 pH 2 1.26 logDOC 1 0.24 pH z logDOC 2 19.26 (R 2 5 0.60). If the changes in soil chemical conditions after afforestation influence the cadmium and copper release rates in a similar way as observed in the flow cell experiments, then the release rate of cadmium will increase exponentially at soil solution pH , 4.5. The inhibition of copper release by DOC observed at pH 3.8 to 5.0 indicates that copper is retained in the soil by interactions with adsorbed organic matter. Copyright

Low-molecular-weight aliphatic carboxylic acids in soil solutions under different vegetations determined by capillary zone electrophoresis

Concentrations of low-molecular-weight aliphatic carboxylic acids in soil solution were determined by a newly developed capillary zone electrophoresis method. Soil solution samples were collected by centrifugation of soil from the A horizon of a Danish, homogeneous, nutrient-rich Hapludalf in adjacent forested and arable plots. The forested plots of 0.5 ha were 33-year old stands of beech (Fagus sylvatica L.), oak (Quercus robur L.), grand fir (Abies grandis Lindl.), and Norway spruce (Picea abies (L.) Karst.), while sugar beet (Beta vulgaris L.) and winter wheat (Triticum aestivum L.) were the agricultural crops this year. High variability in soil solution concentrations of metal cations (Al, Ca, K, Mg, Na), monocarboxylic acids (formic, acetic, lactic, and valeric acids), and di- and tricarboxylic acids (oxalic, malic, succinic, and citric acids) were found within each plot. Despite this short-range within-plot variability, higher concentrations of di- and tricarboxylic acids were found in the forested soils than in the arable soils. The vegetation seemed to favour some monocarboxylic acids, but the total monocarboxylic acid concentrations showed little relation to the vegetation. Probably due to much less soil water in the Norway spruce plot, the low-molecular-weight aliphatic carboxylic acid concentrations in the samples from that plot were much higher than those found in samples from the other plots. Carbon in low-molecular-weight aliphatic carboxylic acids only accounts for a few percent of dissolved organic carbon, and no general relation was found between carbon in low-molecular-weight aliphatic carboxylic acids and dissolved organic carbon, although the correlation between carbon in di- and tricarboxylic acids and dissolved organic carbon was significant.

Effects of a triazole fungicide and a pyrethroid insecticide on the decomposition of leaves in the presence or absence of macroinvertebrate shredders

Previously, laboratory experiments have revealed that freely diluted azole fungicides potentiate the direct toxic effect of pyrethroid insecticides on Daphnia magna. More ecologically relevant exposure scenarios where pesticides are adsorbed have not been addressed. In this study we exposed beech leaves (Fagus sylvatica) to the azole fungicide propiconazole (50 or 500 μg L(-1)), the pyrethroid insecticide alpha-cypermethrin (0.1 or 1 μg L(-1)) or any combination of the two for 3h. Exposed leaves were transferred to aquaria with or without an assemblage of macroinvertebrate shredders, and we studied treatment effects on rates of microbial leaf decomposition, microbial biomass (using C:N ratio as a surrogate measure) and macroinvertebrate shredding activity during 26 days post-exposure. Microbial leaf decomposition rates were significantly reduced in the propiconazole treatments, and the reduction in microbial activity was significantly correlated with loss of microbial biomass (increased C:N ratio). Macroinvertebrate shredding activity was significantly reduced in the alpha-cypermethrin treatments. In addition, the macroinvertebrate assemblage responded to the propiconazole treatments by increasing their consumption of leaf litter with lower microbial biomass, probably to compensate for the reduced nutritional quality of this leaf litter. We found no interaction between the two pesticides on macroinvertebrate shredding activity, using Independent Action as a reference model. In terms of microbial leaf decomposition rates, however, alpha-cypermethrin acted as an antagonist on propiconazole. Based on these results we emphasise the importance of considering indirect effects of pesticides in the risk assessment of surface water ecosystems.

Analysis of selected phytotoxins and mycotoxins in environmental samples

Natural toxins such as phytotoxins and mycotoxins have been studied in food and feed for decades, but little attention has yet been paid to their occurrence in the environment. Because of increasing awareness of the presence and potential relevance of micropollutants in the environment, phytotoxins and mycotoxins should be considered and investigated as part of the chemical cocktail in natural samples. Here, we compile chemical analytical methods to determine important phytotoxins (i.e. phenolic acids, quinones, benzoxazinones, terpenoids, glycoalkaloids, glucosinolates, isothiocyanates, phytosterols, flavonoids, coumestans, lignans, and chalcones) and mycotoxins (i.e. resorcyclic acid lactones, trichothecenes, fumonisins, and aflatoxins) in environmentally relevant matrices such as surface water, waste water-treatment plant influent and effluent, soil, sediment, manure, and sewage sludge. The main problems encountered in many of the reviewed methods were the frequent unavailability of suitable internal standards (especially isotope-labelled analogues) and often absent or fragmentary method optimization and validation.

The effects of tree species and site on the solubility of Cd, Cu, Ni, Pb and Zn in soils

Afforestation of former agricultural land changes soil characteristics such as pH and organic matter content, which may affect heavy metal solubility in the soil. In this study the effects of different tree species on heavy metal solubility were investigated at four 34 years old adjacent stands of beech (Fagus sylvatica L.), grand fir (Abies grandis Lindl.), Norway spruce (Picea abies (L.) Karst.) and oak (Quercus robur L.) planted on former agricultural land at four different sites in Denmark. The sites differ in soil characteristics and represent two texture classes (loamy sand and sandy loam). Soil pH and soil organic matter content was measured in the 16 stands and soil solution was isolated by centrifugation from three depths at four different occasions. Dissolved organic carbon (DOC), pH in the soil solution and the soil solution concentrations (availability) of Cd, Cu, Ni, Pb and Zn were determined. Analysis of variance showed that the tree species affects soil pH and organic matter content in the topsoil, but not in the lower horizons. Norway spruce and grand fir acidify more than beech and oak, and the highest amount of accumulated soil organic matter is in the topsoil under Norway spruce. The effects of tree species on soil solution pH and DOC resemble the effect on soil pH and organic matter content. Grand fir enhances the solubility of Cd and Zn in the topsoil with the lower solubility found under beech and oak and Norway spruce enhances the solubility of Cu, Ni and Pb in the top horizons. The lowest solubility of Ni and Pb is found under beech and oak, whereas the lowest Cu concentrations in the soil solution are found under grand fir. After 34 years of afforestation no effects of tree species on the concentrations of heavy metals in the soil solution from the C-horizons were found. The tree species effect on the concentration of Cd, Cu and Ni in the soil solution depends on the soil characteristics with the higher concentrations found in sandy loam soils, whereas no effect of site on the solution concentration was found for Pb and Zn. It was not possible to find a clear correlation between the soil solution concentrations of heavy metals, pH and DOC concentration.

Pesticide Side Effects in an Agricultural Soil Ecosystem as Measured by amoA Expression Quantification and Bacterial Diversity Changes.

Background and Methods Assessing the effects of pesticide hazards on microbiological processes in the soil is currently based on analyses that provide limited insight into the ongoing processes. This study proposes a more comprehensive approach. The side effects of pesticides may appear as changes in the expression of specific microbial genes or as changes in diversity. To assess the impact of pesticides on gene expression, we focused on the amoA gene, which is involved in ammonia oxidation. We prepared soil microcosms and exposed them to dazomet, mancozeb or no pesticide. We hypothesized that the amount of amoA transcript decreases upon pesticide application, and to test this hypothesis, we used reverse-transcription qPCR. We also hypothesized that bacterial diversity is affected by pesticides. This hypothesis was investigated via 454 sequencing and diversity analysis of the 16S ribosomal RNA and RNA genes, representing the active and total soil bacterial communities, respectively. Results and Conclusion Treatment with dazomet reduced both the bacterial and archaeal amoA transcript numbers by more than two log units and produced long-term effects for more than 28 days. Mancozeb also inhibited the numbers of amoA transcripts, but only transiently. The bacterial and archaeal amoA transcripts were both sensitive bioindicators of pesticide side effects. Additionally, the numbers of bacterial amoA transcripts correlated with nitrate production in N-amended microcosms. Dazomet reduced the total bacterial numbers by one log unit, but the population size was restored after twelve days. The diversity of the active soil bacteria also seemed to be re-established after twelve days. However, the total bacterial diversity as reflected in the 16S ribosomal RNA gene sequences was largely dominated by Firmicutes and Proteobacteria at day twelve, likely reflecting a halt in the growth of early opportunists and the re-establishment of a more diverse population. We observed no effects of mancozeb on diversity.

Ergot alkaloids in rye flour determined by solid-phase cation-exchange and high-pressure liquid chromatography with fluorescence detection

Ergot alkaloids are mycotoxins that are undesirable contaminants of cereal products, particularly rye. A method was developed employing clean-up by cation-exchange solid-phase extraction, separation by high-performance liquid chromatography under alkaline conditions and fluorescence detection. It is capable of separating and quantifying both C8-isomers of ergocornine, α-ergocryptine, ergocristine, ergonovine, and ergotamine. The average recovery was 61%  ± 10% with limits of detection from 0.2 to 1.1 µg kg−1. Twenty-four unknown rye flour samples from Danish mills contained on average 46 µg kg−1 with a maximum content of 234 µg kg−1. The most common ergot alkaloids were ergotamine and α-ergocryptine including their C8-isomers. A total of 54% of the ergot alkaloids were detected as C(8)-S isomers.