Liquid extraction of low molecular mass organic acids and hydroxamate siderophores from boreal forest soil

Low molecular mass organic acids (LMMOAs) and hydroxamate siderophores (HS) are molecules secreted by microbes and have previously been found in soil solution and in cultures. Mycorrhizal fungi are suggested to be involved in the nutrient uptake processes of trees and weathering of minerals. In this study soil samples taken from the O and E horizons of a podzol were extracted with 10 mM potassium phosphate buffer at pH 7.2. Variable parameters included addition of methanol to the extraction buffer and the use of ultrasonication or rotary shaking during extraction. LMMOAs and HS content of the soil extracts were determined. Analysis of soil extracts were carried out by liquid chromatography mass spectrometry (LC–MS) and the extraction results compared to results for soil solution samples obtained by centrifugation of the soils sampled. The extraction yields were significantly increased by addition of methanol to the extraction buffer, especially for the O horizon samples. Rotary shaking of the samples for 90 min gave slightly higher yields than ultrasonication for 15 min but the reduction in extraction time makes ultrasonication an attractive option. Of the HSs determined, ferricrocin was found in all samples. Optimal extraction conditions showed citric acid and isocitric acid to be the most abundant organic acids in the O and E horizons, respectively.     

Comparison of two alkaline treatments in the extraction of organic compounds associated with water repellency in soil under Pinus taeda

Isopropanol:NH₃ and Methanol:KOH (saponification) alkaline treatments are usually applied in the study of soil organic matter. The first is used in studies of soil water repellency, and the latter in the extraction of ester-bound lipids from soil. In this study, isopropanol:NH₃ and methanol:KOH treatments were applied separately in a solvent extracted repellent soil, in order to compare their efficiency in the extraction of water repellent compounds. The soil sample was taken from a site under a 16 year old Pinus taeda stand. The amount and class of organic compounds released by each treatment were compared using gas chromatography-mass spectrometry (GC-MS). Both treatments resulted in wettable soil after alkaline extraction. In general, alkaline treatments yielded extracts with the same class of organic compounds. Alkanoic acids, α,ω-alkanedioic acids, hydroxyalkanoic acids, aromatic compounds, and alkanols were identified, indicating the preservation of suberin and cutin biopolyester in the soil. Large differences were observed in the amounts of ω-hydroxyalkanoic acids, as well as in the quantity and distribution of dihydroxyalkanoic and trihydroxyalkanoic acids. In contrast to methanol:KOH, isopropanol:NH₃ was not efficient in the extraction of whole aliphatic biopolyesters, mainly pine cutin-related products. Methanol:KOH was more effective in hydrolysis. The presence of biopolyesters in water repellent soil under the P. taeda stand seems to play an important role in water repellency.     

Buffer capacities of podzolic and peat gleyic podzolic soils to sulfuric and nitric acids

Soil samples from the main genetic horizons of pale podzolic and peat gleyic podzolic soils from the Central Forest Reserve were subjected to a continuous potentiometric titration by sulfuric and nitric acids. The sulfate sorption capacity was determined in soil mineral horizons. The buffer capacity of mineral horizons of both soils to sulfuric acid was found to be higher than that to the nitric acid. This is explained by the sorption of sulfates via the mechanism of ligand exchange with the release of hydroxyl groups from the surfaces of Fe and Al hydroxide particles and edge faces of clay crystallites. The buffer capacity of organic horizons of the pale podzolic soil to sulfuric acid proved to be higher than that to nitric acid; in organic horizons of the peat gleyic podzolic soil, the buffer capacity to sulfuric acid was lower than that to nitric acid. The reasons for this phenomenon have yet to be investigated.     

Pre-lysis washing improves DNA extraction from a forest soil

A pre-lysis buffer washing procedure was introduced to DNA extraction from a forest soil with high organic matter and iron oxide contents. Sodium phosphate of 0.1 M (pH 7.5) was used as a buffer to wash soil samples when subsequent lysis buffer was phosphate, and 20 mM EDTA (pH 7.5) was used when subsequent lysis buffer included EDTA. Initial experiments were not successful because the DNA extracts could not be amplified by polymerase chain reaction (PCR). The consideration of introducing a pre-lysis washing procedure was based on the idea that the washing should promote soil dispersion and homogeneity, decrease DNA adsorption by soil components (e.g. iron oxides), and remove covalent cations and those easily-dissolving organic compounds from the soil samples. Results revealed that humic substance content decreased by 31%, but DNA yield increased by 24% in the DNA extracts of the pre-lysis washing procedures, compared to the non-washing procedures. DNA extracted by the pre-washing procedure needed less purification for subsequent 18S and 16S rDNA PCR amplifications. It was recommended that the pre-lysis buffer washing should be used for DNA extraction from those difficult environmental samples, such as the forest soil with high contents of organic matter and iron oxides.     

Loss of low molecular weight dissolved organic carbon (DOC) and nitrogen (DON) in H2O and 0.5 M K2SO4 soil extracts

Soil extracts are routinely used to quantify dissolved organic nutrient concentrations in soil. Here we studied the loss and transformation of low molecular weight (LMW) components of DOC (¹⁴C-glucose, 1 and 100 μM) and DON (¹⁴C-amino acid mixture, 1 and 100 μM) during extraction of soil (0-6 h) with either distilled water or 0.5 M K₂SO₄. The extractions were performed at 20 °C, at 4 °C, or in the presence of an inhibitor of microbial activity (HgCl₂ and Na-azide). We showed that both glucose and amino acids became progressively lost from solution with increasing shaking time. The greatest loss was observed in H₂O extracts at 1 μM for both substances (>90% loss after 15 min). Lower temperature (4 °C) and presence of K₂SO₄ both resulted in reduced loss rates. The presence of microbial inhibitors effectively eliminated the loss of glucose and amino acids. We conclude that microbial transformation of LMW-DOC and DON during H₂O or K₂SO₄ extraction of soil may affect the estimation of their concentrations in soil. This finding has significant implications for methods that rely on chemical extractions to estimate LMW-C components of DOC and DON.     

Low molecular weight organic acid adsorption in forest soils: effects on soil solution concentrations and biodegradation rates

Low molecular weight (LMW) organic acids are believed to play a key role in many rhizosphere and pedogenic processes; However, their efficiency is likely to depend on their susceptibility to sorption and biodegradation. The sorption characteristics of three organic acids (citrate, oxalate and acetate) and phosphate were examined over the concentration range 0-1000 μM in three coniferous forest soil profiles. Sorption to the soil's solid phase could be adequately described by the Langmuir equation with sorption capacity following the horizon series: B>C>E>O. The strength of anion sorption followed the series: phosphate>oxalate≥citrate?acetate. Calculations indicated that between 50 and 95% (O and E horizons) and >93% (B horizons) of these LMW organic acids entering the soil will become sorbed to the solid phase. The amount of organic acids predicted to be present on the solid phase at typical soil solution concentrations ranged from <1 to 1100 nmol g⁻¹ yielding adsorbed-to-solution ratios (adsorption coefficients) of between <0.1 and 3100. In the case of citrate, sorption to the solid phase significantly reduced its biodegradation potential by 35-99% depending upon the degree and type of sorption surface. The findings of this work are discussed in the context of the quantitative effects of adsorption on organic acids, their ecological functions and role in soil forming processes.     

Extraction of bacteria from soil: Efficiency of shaking or ultrasonication as indicated by direct counts and autoradiography

Blending, shaking with a mechanical wrist-action shaker or ultrasonication were compared for removing bacteria from a sandy soil and a silt loam soil. Bacteria were counted microscopically after staining with acridine orange. There was no significant difference in the numbers of bacteria released from the sandy soil by the different treatments but ultrasonication extracted more bacteria from the silt loam. Highest recoveries of bacteria were obtained with a combination of Tris buffer as extractant, and 25% Ringer's solution as diluent. Ultrasonication of an irradiated sandy soil containing added Bacillus cereus removed 97% of the bacteria from the sand grains, but lysed some cells. Members of the indigenous soil population were more firmly held by the soil than added bacteria. Optimum conditions for recovering soil bacteria by ultrasonication were either 10.5 μm for 30s or 5 μm for 2 min. With the natural population, the numbers of bacteria and the proportion of metabolizing bacteria increased a little after ultrasonication as shown by labelling cells with ³H-glucose and detecting uptake by autoradiography. However, three isolates of bacteria tested showed varying amounts of injury after ultrasonication, with 5 μm for 2 min causing more damage than 10.5 μm for 30s. Ultrasonication was a more efficient means of extracting the natural bacterial population of soil than shaking but the time and amplitude must be adjusted to avoid injury to cultured bacterial cells.     

Modelling low molecular weight organic acid dynamics in forest soils

Low molecular weight organic acids such as citrate and oxalate have been hypothesized to play a key role in rhizosphere ecology and pedogenesis. A mathematical site-specific model, DYNLOW, was constructed to describe the temporal and spatial dynamics of these organic acids in coniferous forest soils using the modelling software STELLA^®. Experimentally derived values for biodegradation, adsorption, and daily values of soil temperature, moisture and hydrological flow were used to parameterize the model. The model describes the dynamics and downward movement of oxalate and citrate through the horizons (O, AE, E, Bhs, Bs) of three podzolic soil profiles in Sweden. After calibration, the model predicted average soil solution organic acid concentrations ranging from <1 to 90 μM, which was in agreement with experimental measurements (<1 to 116 μM). The model results indicated that microbial degradation of organic acids was in quantitative terms the biggest process regulating soil solution concentrations. Primary production rates of organic acid in the soil were predicted to be high (<1 to 1250 nmol g⁻¹ soil d⁻¹) in comparison to the amount present at steady state in the soil solution pool (<0.1 to 240 nmol g⁻¹ soil). The downward transfer of organic acids between soil horizons due to mass flow was predicted to be a small flux (<0.1 to 3% of the total loss) compared to that lost by microbial biodegradation. The model predicted that the amount of basal soil respiration that could be attributable to the microbial turnover of organic acids was on average 19±22% of the basal CO₂ production across all sites and horizons for citrate and 7±7% for oxalate. The model results are discussed in the context of pedogenesis, forest soil respiration and organic matter production.     

Acidity in organic horizons of arctic soils on the Barents Sea coast

Parameters of water and KCl extracts from organic horizons of arctic soils on the coast of Khaipudyr Bay of the Barents Sea, in which the values of pH are 3.8–4.3 and 2.7–3.5, respectively, have been compared. It has been found that the content of water-extractable organic carbon is 0.2–0.5 g/dm³; the contents of low-molecular-weight carbohydrates and acids are 1–24 and 6 mg/dm³, respectively, and the content of Fe³⁺ ions is 1–4 mmol/dm³. The increase in the ionic strength of solution reduces the extraction efficiency of total organic carbon in 1.2 times and that of acids and carbohydrates in 3 times on the average at the simultaneous increase in the content of iron ions in the liquid phase in 7–15 times. It has been shown that organic acids and iron compounds are the main sources of acidity in salt extracts from organic soil horizons. The low contents of Ca²⁺ and Mg²⁺, which participate in the neutralization of acids, favor the high acidity of the studied horizons.     

Comparison of methods for extraction of organic N monomers from soil microbial biomass

One way of investigating the function of soil is via the pool of low molecular weight organic compounds in the soil microbial biomass. This is because low molecular weight organic compounds have key roles in metabolism of soil microbes, can function in osmotic adjustment and other stress responses, and are intermediates in the breakdown of polymers to inorganic nutrients. Methods for measuring low molecular weight microbial metabolites in soil rely upon extracting total metabolites and then subtracting the contribution from metabolites in the soil extracellular matrix (i.e. microbial = total − extracellular). Recent studies have tested methods for extracting organic N monomers from the extracellular matrix of soil, but there has not been similar testing of methods for extracting total organic N monomers. The aims of this study were to examine methods for extracting total organic N monomers by a) contrasting chloroform gas fumigation with chloroform direct extraction, and b) examining whether it is possible to extract soil with two methods that combine quenching of metabolic activity with extraction, namely cold methanol/chloroform/water and hot aqueous ethanol. To evaluate methods, organic N compounds were extracted from soil and then capillary electrophoresis–mass spectrometry identified and quantified 42 organic N monomers including amino acids, quaternary ammonium compounds, nucleobases and nucleosides, amines and polyamines. Absolute concentrations of 32 out of the 42 quantified organic N monomers were significantly different between soil extracted by chloroform gas fumigation and chloroform direct extraction. These differences were probably a function of gains and losses of compounds due to oxidation, hydrolysis and deamidation during the two-day chloroform gas fumigation. Cold methanol/chloroform/water yielded large amounts of the extremely labile compound ergothioneine, probably because the extraction method rapidly quenched metabolic activity. The primary limitation of extraction with methanol/chloroform/water is that it was ineffective at extracting strongly cationic compounds (e.g. polyamines). Extraction with hot aqueous ethanol was unsuccessful with soil presumably because soil microbes are difficult to lyse. It is recommended that future studies examining organic N monomers in soil microbial biomass use chloroform direct extraction or cold methanol/chloroform/water rather than chloroform gas fumigation.     

Microbially available carbon in buried riparian soils in a glaciated landscape

Buried horizons and lenses in riparian soil profiles harbor large amounts of carbon relative to the surrounding soil horizons. Because these buried soil horizons, as well as deep surface horizons, frequently lie beneath the water table, their impact on nitrogen transport across the terrestrial–aquatic interface depends upon their frequency and spatial distribution, and upon the lability of associated organic matter. We collected samples of 51 soil horizons from 14 riparian zones Rhode Island, USA, where soil profiles are characterized by glacial outwash and alluvial deposits. These soil samples came from as deep as 2 m and ranged in carbon content from <1% to 44% in a buried O horizon 54–74 cm deep. We used these samples to: (1) determine the extent to which carbon in buried horizons, and deep surface horizons, is potentially microbially available; (2) identify spatial patterns of carbon mineralization associated with surface and buried horizons; and (3) evaluate likely relationships between soil horizon types, chemical characteristics and carbon mineralization. Carbon mineralization rates associated with buried horizons during anaerobic incubations ranged from 0.0001 to 0.0175 μmol C kg soil^?1 s^?1 and correlated positively with microbial biomass (R=0.89, P<0.0001, n=21). Excluding surface O horizons from the analysis, carbon mineralization varied systematically with horizon type (surface A, buried A, buried O, lenses, A/C, B, C) (P<0.05) but not with depth or depth x horizon interaction (overall R²=0.59, P<0.0005, n=47). In contrast to this result and to most published data sets, ¹³C-to-¹²C and ¹⁵N-to-¹⁴N ratios of organic matter declined with depth (¹³C?26.9 to ?29.3 per mil, ¹⁵N+5.6 to ?0.8 per mil). The absence of a relationship between horizon depth and C availability suggests that carbon availability in these buried horizons may be determined by the abundance and quality of organic matter at the time of horizon formation or burial, rather than by duration since burial, and implies that subsurface microbial activity is largely disconnected from surface ecosystems. Our results contribute to the emerging view that buried horizons harbor microbially available C in quantities relevant to ecosystem processes, and suggest that buried C-rich soil horizons need to be incorporated into assessments of the depth of the biologically active zone in near-stream subsurface soils.     

Comparison of 0.1N sodium hydroxide with 0.1M sodium pyrophosphate in the extraction of soil organic matter from various soil horizons

Abstract

Soil samples representing the O, A, B, and C soil horizons from soil organic matter (SOM) studies were selected to study the relative effectiveness of 0.1N sodium hydroxide (NaOH) and 0.1M sodium pyrophosphate (NaPyr) in extracting organic carbon (OC). Replicate samples were extracted with each extractant in a ratio of about 1:144 and successive extractions were performed for each soil. Results indicated the importance of successive extractions for more complete removal of extractable OC. A single extraction removed an average of only 68%, 78%, 86%, and 60% of the OC extracted with four successive extractions for the O, A, B, and C horizons, respectively. The C horizons were lowest in OC and the slowest to release OC in the extraction process. Organic C was solubilized from the B horizons most quickly with an average of 95% of the successively extractable OC removed with only two extractions. The extractability of the soil TOC was highest in the Bhs and then the Bw, C, A, and O horizons at 92% and 42%, 46%, 38%, and 3 6%, respectively. The NaOH and NaPyr were nearly equal in extracting OC from the Bhs horizons. The NaOH extracted more OC than the NaPyr at 53%, 55%, 29%, and 47% more in the O, A, Bw, and C horizon samples, respectively. These results stress the importance of considering the soil horizon type and the use of NaOH in successive extraction for maximum removal of OC in soil studies.     

Ein neues Verfahren zur Extraktion von organischen Stoffen aus Böden mit überkritischen Gasen

A new procedure for the extraction of organic matter from soils by supercritical gases Organic solvents beyond critical temperature and pressure have excellent properties for solving polymere organic substances. For soil samples yields of organic material with low ash content are much higher than those obtained by the common NaOH/HCl-extraction. Fractionation of the samples is easily achieved by a simple change of solvent bottles. In comparison to the original material oxygen contents are lower, due to a thermal or solvolytical split off of methanol or water. Extraction with supercritical gases – in addition to oxidative degradation –may be regarded as a useful method to enlighten the structure of humic acids. The effective extraction of firmly bound pesticides in soil is another point of interest.     

Aggregation of clay in six New Zealand soil types as measured by disaggregation procedures

Measurements were made of the yields and carbon concentrations of clay fractions obtained by a number of disaggregation procedures from samples of a range of New Zealand soils. Disaggregation was carried out in water by: minimal shaking by hand; prolonged mechanical shaking; shaking with sodium resin; ultrasonication; acetylacetone treatment; periodate–borate treatment; peroxidation; and peroxidation followed by citrate-dithionite-bicarbonate treatment. There was virtually no clay yielded by any of the six samples when gently suspended in water. Either a combined hydrogen peroxide and sodium citrate-dithionite-bicarbonate treatment or an exhaustive (10× repeated) hydrogen peroxide treatment produced the highest yield of clay except where ultrasonication of the field moist soil sample brought about strong disaggregation. Considerable physical disruption of primary particles could occur on ultrasonication. Air-drying had a variety of effects. In one notable case with a soil having a highly smectitic clay mineralogy, air-drying led to considerable resistance of the sample against disaggregation into clay. Exhaustive peroxidation was usually highly dispersive but it suppressed clay dispersion in samples of two soils from volcanic ash with allophanic clay mineralogies.There was a wide range between samples of the different soils in the concentrations and amounts of carbon in their clay fractions. A high concentration of carbon was present in clay fractions of all samples after even the most severe chemical treatment, with those of the allophanic soils having the highest concentrations of carbon after these treatments.     

Fractionation of dissolved organic carbon in soil water: Effects of extraction and storage methods

Abstract

We measured the concentration and composition (sensu Leenheer, 1981) of dissolved organic carbon (DOC) in lysimeter solutions from the forest floor of a spruce stand in Maine and in laboratory extracts of organic (Oa horizon) and mineral soils collected from various forests in Maine, New Hampshire, and Vermont. All soils were acid Spodosols developed from glacial till. The effects of different storage, extraction and filtration methods were compared. Extracts from Oa horizons stored fresh at 3°C contained a larger fraction of hydrophobic neutrals than lysimeter forest floor solutions (31 and 4% of DOC in stored and lysimeter solutions, respectively), whereas extracts from Oa horizons which had been extracted, incubated at 10–15°C, and extracted again had DOC compositions similar to that in lysimeter solutions. Mechanical vacuum and batch extractions of Oa horizons yielded DOC similar in concentration and composition if the extracts were filtered through glass fiber filters. Nylon membrane filters, however, removed more hydrophobic acids from batch extracts. Dissolved organic carbon extracted from frozen, air‐dry, and oven‐dry Oa and Bh horizons was relatively rich in hydrophilic bases and neutrals and was similar to that released after chloroform fumigation, indicating that common soil‐storage methods disrupt microbial biomass.     

Organic matter dynamics in a temperate forest soil following enhanced drying

Climate models predict an increase in global surface temperature and a change in precipitation intensity during this century. For Europe, extended drought periods followed by heavy rainfall are expected. The consequences for soil organic matter (SOM) dynamics are poorly understood. In this study, we investigated the effect of changing soil moisture regime on SOM quality under field conditions. For this purpose, a throughfall exclusion (TE) experiment was conducted in the summers 2006 and 2007 on a Haplic Podzol under a 140 years old Norway spruce stand using a roof installation followed by re-wetting compared to non-manipulated control plots. Total organic carbon, lignin (stable carbon pool), plant and microbial sugars (labile carbon pool) and microbial biomass (phospholipid fatty acids) were determined before, during and after the experiment in the L, O, A and B horizons. No significant treatment effects could be observed for SOM quantity. Amounts of lignin and soil microbial biomass were also not affected by the moisture regime but structure of soil microbial community. In the L and organic layers, gram + bacteria and actinomycetes were reduced during water stress, while gram- bacteria, fungi and protozoa increased during drought. Warmer and drier weather led to a dominance of fungi while a cooler and moister regime favoured bacteria, at least in the L horizon. An increasing PLFA (cy17:0 + cy19:0)/(16:1ω7c + 18:1ω7c) ratio in the O layer and A horizon suggests that the microbes suffered from water stress in these horizons. This agrees with a decreasing contribution of microbial sugars to SOM with decreasing water content in the O and A horizons. Although the original plant material exhibited increasing plant sugar content with increasing dryness, the contribution of the plant sugars to total soil organic carbon (SOC) generally decreased with decreasing water content. Physical-chemical changes of soil structure can theoretically change the sugar extractability from soils and/or chemical changes of sugars structure can probably affect the analysis. Therefore, chemical alteration and stabilization could be responsible for sugar decrease in soil with increasing dryness explaining the contrast compared to the original plant material.     

Quantification of Escherichia coli O157:H7 in soils using an inhibitor-resistant NanoGene assay

Humic acids are ubiquitous and abundant in terrestrial environments; therefore, they are often co-extracted with nucleic acids and interfere with quantitative PCR (qPCR) assays. In this study a recently developed NanoGene assay that is resistant to interference by humic acids was evaluated for gene detection in soil samples. The NanoGene assay utilizes a combination of magnetic beads, dual quantum dots labels, and DNA hybridization in solution. Seven soil samples containing different amounts of organic matter were tested to compare NanoGene and qPCR assays for their respective ability to detect a bacterial pathogen. We spiked the soils with Escherichia coli O157:H7, extracted genomic DNA, and conducted NanoGene and qPCR assays targeting the E. coli O157:H7-specific eaeA gene. To prevent the inhibition of PCR that is common when using DNA extracted from soils, we used a range of template DNA concentrations and BSA addition in the qPCR assay. Compared to the qPCR assay the NanoGene assay was significantly more resistant to the inhibitory effect of humic acids, successfully quantifying the eaeA gene within a linear (R² = 0.99) range of 10⁵ through 10⁸ CFU/g soil for all seven soil samples tested. In contrast, the qPCR assay was significantly inhibited using the same template DNA isolated from soils containing a range of organic content (2.0%–12%). Interestingly, the qPCR assay was still inhibited despite additional purification steps, suggesting that humic acids were still associated with DNA at a level that was inhibitory to qPCR. This study demonstrated that the NanoGene assay is suitable for quantitative gene detection in diverse soil types and is not susceptible to inhibition by humic acids and other organic compounds that commonly lead to false negative results in qPCR assays.     

Available organic nitrogen in temperate, subtropical, and tropical soils extracted with different solutions

Extraction of organic N by chemical solutions has been used to assess the amount of available N in soil. We tested the efficiency of several solutions in extracting organic N from tropical, subtropical and temperate soils. A conventional 0.067 M phosphate buffer successfully extracted organic N from all 23 soils examined. High-performance size exclusion chromatograms showed a single peak at about 7,800 Da for all phosphate buffer extracts irrespective of soil types. The peak area correlated with the organic N concentration of extracts. Tropical soils had lower retention of organic N than other soils according to the conventional and sequential extraction with phosphate buffer. Organic N extracted with sulfuric acid was significantly (P < 0.001) correlated with the amount of extracted Fe, suggesting that Fe might play a role in the retention of organic N in soil.     

Organic acid induced release of nutrients from metal-stabilized soil organic matter – The unbutton model

Processes of soil organic matter (SOM) stabilization and the reverse, destabilization of SOM resulting in subsequent release and mobilization of nutrients from SOM, remain largely unresolved. The perception of SOM as supramolecular aggregates built of low molecular mass biomolecules is currently emerging. Polyvalent metal cations contribute to SOM tertiary structure by bridging functional groups of such molecules (Simpson et al., 2002). The strong bond to metals protects high quality organic material from being immediately accessed and decomposed. Here we propose a three-step process by which low molecular mass organic acids (LMMOAs) and hydrolytic enzymes act in series to destabilize SOM supramolecules to release organic nitrogen (N) and phosphorus (P) for local hyphal and root uptake. Complexation of the stabilizing metals by fungal-released LMMOA gives fungal-root consortia direct access to organic substrates of good quality. Because of their small sizes and carboxyl group configuration, citric and oxalic acids are the most effective LMMOAs forming stable complexes with the main SOM bridging metals Ca and Al in SOM. Citrate, forming particularly strong complexes with the trivalent cations Al and Fe, is dominant in soil solutions of low-productive highly acidic boreal forest soils where mycorrhizal associations with roots are formed predominantly by fungi with hydrophobic hyphal surfaces. In these systems mycelia participate in the formation of N-containing SOM with a significant contribution from strong Al bridges. In less acidic soils of temperate forests, including calcareous influenced soils, SOM is stabilized predominantly by Ca bridges. In such systems mycorrhizal fungi with more hydrophilic surfaces dominate, and oxalic acid, forming strong bidentate complexes with Ca, is the most common LMMOA exuded. A plant-fungus driven biotic mechanism at the supramolecular aggregate level (10³–10⁵ Da) resolves micro-spatial priming of SOM, where the destabilization step is prerequisite for subsequent release of nutrients.     

The effect of low-molecular-weight organic acids and inorganic phosphorus concentration on the determination of soil phosphorus by the molybdenum blue reaction

Low-molecular-weight organic acids play an important role in the mobilization of soil phosphorus (P). The molybdate blue colorimetric method, based on the formation of P molybdenum blue compound, is commonly used for analysis of phosphate in soil and environmental samples. However, some organic acids can act as a ligand to bond with molybdenum acid, which subsequently interfere with the colorimetric reaction. The recoveries of P were inhibited by the addition of oxalic (>2 mM) and citric acids (>3 mM) both in standard P solutions and soil extracts, but formic and maleic acids did not interfere with the P determination. The inhibition of oxalic and citric acids on P recovery remained even at higher level of P (up to 100 μg P 100 ml⁻¹) though such interferences decreased by increasing P concentration. Comparison between oxalic and citric acids revealed the more pronounced interference by the addition of oxalic acid. The results suggested that the interference of organic acids with P determination is related to the types of organic acids and the ratio of organic acid ligands to P anions in the solutions. Thus, analysis of P using the molybdate blue colorimetric method should be undertaken cautiously in the presence of relative strong ligands like oxalic and citric acids.