A new method for measuring microbial biomass nitrogen in soils: Direct extraction after toluene treatment

Abstract

Phenolic compounds occurring naturally are reducing agents which react with hydrous Fe oxide and Mn oxides (Lehmann et al. 1987). Diphenols such as hydroquinone, resorcinol, and catechol were oxidatively darkened by the presence of hydrous Fe oxide though the degree of darkening was much less pronounced than that by Mn oxides (Shindo and Huang 1984). Okazaki et al. (1976) suggested that polyphenols are one of the materials responsible for the dissolution of Mn in a paddy soil under reducing conditions.     

Selective digestion of the peptide and polysaccharide components of synthetic humic acids by the humivorous larva of Pachnoda ephippiata (Coleoptera: Scarabaeidae)

In order to identify the potential nutrient and energy sources of humivorous beetle larvae, we carried out feeding trials with soil supplemented with specifically ¹⁴C-labeled model humic acids synthesized by peroxidase-initiated radical polymerization, using the cetoniid beetle Pachnoda ephippiata (Coleoptera: Scarabaeidae) as a model organism. Ingestion of soil by the larvae significantly increased the mineralization of humic acids labeled in their peptide (HA-*peptide) or polysaccharide components (HA-*peptidoglycan and HA-*chitin), whereas the mineralization of humic acids labeled in the aromatic components (HA-*catechol) did not increase significantly. Mineralization was accompanied by a reduction of residual radiolabel in the acid-soluble fraction and an increase in the humic acid and humin fractions of the fecal pellets. During the gut passage, the residual label in peptide or polysaccharide components was transformed into acid-soluble products, especially in the alkaline midgut. High-performance gel-permeation chromatography demonstrated that the changes in solubility were accompanied by large changes in the molecular weight of the residual material. The amount of radiolabel derived from the peptide and polysaccharide components recovered from the larval body and hemolymph was significantly higher than that derived from the aromatic component, which supports the hypothesis that humivorous beetle larvae selectively digest the peptide and polysaccharide components of humic substances, whereas the aromatic components of humic substances are not an important source of nutrients and energy. This is also the first experimental evidence that also chitin and peptidoglycan, the major structural polymers in fungal and bacterial biomass, can be protected from microbial degradation in soil by a copolymerization with phenols and might contribute substantially to the refractory nitrogen pool in soil organic matter.     

Preparation and characterization of model humic polymers containing organic phosphorus

The nature of organic P in soil organic matter was studied by evaluating the incorporation of serine, phosphoserine, ethanolamine, phosphoethanolamine and glycerophosphate into model humic polymers prepared by chemical oxidation of polyphenols. Elemental and functional group analysis indicated that the composition of model humic polymers ranged as follows: organic C, 50.6–56.8%; total acidity, 7.86–11.87m-equiv g^?1; carboxyl, 1.42–2.00 m-equiv g^?1; total hydroxyl, 6.79-10.0 m-equiv g^?1; ash, 6.4–13.9%; E₄/E₆ ratio, 5.34–6.19; organic N, 0.70–1.65% and organic P, 0.254–0.942%. These values are within the ranges reported for soil humic substances. The only non-phenolic compounds incorporated into model humic polymers were those containing free amino groups. The P content of model polymers was not increased by the presence of KH₂PO₄, glycerophosphate, serine or ethanolamine whereas phosphoserine and phosphoethanolamine resulted in model polymers containing 0.254 and 0.942% P, respectively. Further characterization studies of the model polymer containing phosphoethanolamine (HA-PE) showed that most of the C (83.2%), N (79.8%) and P (75.3%) was in the humic acid fraction. Gel filtration of HA-PE showed that 0.5% of the polymer was present in high molecular weight (mol. wt) components (mol. wt > 100,000) and 74.8% of the polymer was in two components of mol. wt 10,000–50,000. The majority of the organic P in HA-PE was associated with the medium molecular weight fractions (79.2%) while 16.8% of the P was associated with materials possessing mol. wt < 10,000. Attempts to demonstrate the presence of organic P functional groups contained in HA-PE by infrared spectroscopy was limited by the relatively small amounts of organic P incorporated into the model humic polymers. The results obtained show that a portion of the unidentified organic P in soil humic substances may arise from the incorporation of organic compounds containing both amino and phosphate ester functional groups during oxidative polymerization of polyphenols.     

Nitrogenous constituents in nest soils of harvester ants (messov ebeninus) and their influence on plant growth

Abstract

As a corollary of a detailed study on the ecology of the Harvester ants in soils of semi‐arid natural pastures, samples from a loessial Arid Brown soil were analyzed for the content of nitrogenous constituents (humic compounds, amino acids, nitrates, ammonia) as well as phenols and carbohydrates and the nutrient status of the soil.

While there was a significant increase in fulvic and humic acids in the ant nests, the simultaneous decrease in amino acids ‐ in relation to the surrounding soil ‐ indicates that the humic substances have been synthesized from amino acids and the abundantly present phenols and possibly carbohydrates.

The much higher concentration of nitrogen compounds in the ant mounds supports the idea that the soil mixing activity of the Harvester ants enhances microbiological processes and in turn the fertility of the mounds, as evident in the better growth of the pasture plants.     

The sodium-amalgam reduction of soil and fungal humic substances

Two soil humic acids and a “humic acid” synthesized in the laboratory by Stachybotrys chartarum were reduced with Na-amalgam. The reduction products were methylated, separated by preparative gas chromatography and identified by matching their mass and micro-infrared spectra with those of authentic specimsens.Yields of reduction products identified ranged from 2.7 to 4.2% of the initial weights of the humic materials. Major products identified were N-methyl-benzylsulfonamide, methylated phenolic acids, aromatic aldehydes and C₆C₂ - and C₆C₃ - compounds with 0 in the side chains. Since the Na-amalgam reduction of both soil and fungal humic materials produces the same or similar compounds, the method provides little information on the origin of these compounds, that is, whether they are lignin- or flavonoid-derived or synthesized by microorganisms. Compared with oxidative degradation methods, Na-amalgam reduction appears to be inefficient and tells little about the chemical structure of humic acid polymers.     

Elemental composition and structural features of humic substances in young podzols developed on sand quarry dumps

Data were analyzed on the elemental composition of humic and fulvic acids isolated from organic and mineral horizons of different-aged podzols developed on sand quarry dumps in Leningrad oblast. It was shown that the humic substances from the young podzols significantly differed from those isolated from the mature soils. The humic acids (HAs) of the young podzols composed a more homogeneous group of natural compounds compared to the fulvic acids (FAs). The HAs isolated from the organic, podzolic, and illuvial horizons of the embryopodzols significantly differed in their structure. The litter HAs were characterized by the minimum aromaticity and the maximum content of aliphatic components. The HAs from the illuvial horizons contained the maximum portion of aromatic components; a significant portion of carboxyls, ethers, polysaccharides, carbohydrates, and amino acids; and the minimum portion of aliphatic components. The HAs from the podzolic horizons occupied an intermediate position between the humic substances of the organic and illuvial horizons. It was shown that Mendeleev’s formula was more suitable than Aliev’s formula for calculating the calorific values of the HAs because of their structural features. Based on the particular features of the humic substances studied, they were classified as prohumic substances.     

Crystallization of single and binary iron‐ and aluminum hydroxides affect phosphorus desorption

In acidic soils, phosphorus availability is affected by its strong affinity for mineral surfaces, especially Fe‐ and Al‐hydroxides. Plant roots have developed adaptive strategies to enhance the availability of phosphorus, including producing and exuding low molecular weight organic acids with a high affinity for phosphorus that competes with high molecular weight organic ligands formed during humification and mineralization. The aim of this study was to characterize the kinetics and mechanism of phosphorus desorption from Fe‐ and Al‐hydroxides of variable crystallinity, as well as binary Fe:Al‐hydroxide mixtures. Long‐term desorption experiments (56 days) were conducted with CaCl₂, CaSO₄, citric acid, and humic acid as competitive sorptives. CaCl₂ and CaSO₄ were selected as general inorganic sorptives and citric and humic acids were selected as organic ligands produced by organisms in the rhizosphere or following humification. The cumulative phosphorus desorption increased following the order CaCl₂ < CaSO₄ < humic acid < citric acid. Amorphous ferrihydrite and Fe‐rich Fe:Al‐hydroxides exhibited much less desorption when exposed to inorganic solutions than the crystalline and Al‐rich Fe:Al‐hydroxide mixtures. Models of the desorption data suggest phosphorus desorption with citric acid is diffusion‐controlled for ferrihydrite and Fe‐rich amorphous Fe:Al‐hydroxides. When humic acid was the sorptive, metal‐organic complexes accumulated in the solution. The results suggest organic compounds, especially citric acid, are more important for liberating phosphorus from Fe‐ and Al‐minerals than inorganic ions present in the soil solution.     

Enrichment of natural 15N abundance in frankia-infected nodules

Abstract

The enrichment of ¹⁵N in the nodules of some N₂-fixing leguminous plants is an interesting finding (Shearer et al. 1982). The extent of ¹⁵N enrichment differed depending on the plant species (Shearer et al. 1982; Yoneyama 1987) and bacterial strains (Steele et al. 1983), and in soybeans it was apparently related to the nitrogen fixation efficiency (Shearer et al. 1984)     

The chemistry of soil organic nitrogen: a review

1. From the data presented herein it is possible to deduce the following distribution of total N in humic substances and soils: proteinaceous materials (proteins, peptides, and amino acids) – ca. 40%; amino sugars – 5–6%; heterocyclic N compounds (including purines and pyrimidines) – ca. 35%; NH₃–19%; approximately 1/4 of the NH₃ is fixed NH₄ ⁺. Thus, proteinaceous materials and heterocyclics appear to be major soil N components. 2. Natural ¹⁵N abundance levels in soils and humic materials are so low that direct analysis by ¹⁵N NMR is very difficult or impossible. To overcome this difficulty, the soil or humic material is incubated with ¹⁵N-enriched fertilizer. Even incubation in the laboratory for up to 630 days does not produce the same types of ¹⁵N compounds that are formed in soils and humic materials over hundreds or thousands of years. For example, very few ¹⁵N-labelled heterocyclics are detected by ¹⁵N NMR. Does this mean that heterocyclics are not present? Or are the heterocyclics that are present not labelled under these experimental conditions and therefore not detected by the ¹⁵N NMR spectrometer ? Another possibility is that a large number of N heterocyclics occur in soils, but each type occurs in very low concentrations. Until the sensitivity is improved, ¹⁵N NMR will not provide results that can be compared with data obtained from the same soil and humic material samples by chemical methods and mass spectroscopy. 3. What is most important with respect to agricultural is that all major N forms in soils are available to organisms and are sources of NH₃ or NH₄ ⁺ for plant roots and microbes. Naturally, some of the NH₃ will enter the N cycle. 4. From chemical and pyrolysis-mass spectrometric analyses it appears that N heterocylics are significant components of the SOM, rather than degradation products of other molecules due to pyrolysis. The arguments in favor of N heterocyclics as genuine SOM components are the following: a) Some N-heterocyclics originate from biological precursors of SOM, such as proteinaceous materials, carbohydrates, chlorophyll, nucleic acids, and alkaloids, which enter the soil system as plant residues or remains of animals. b) In aquatic humic substances and dissolved organic matter (DOM) at considerably lower pyrolysis temperatures (200 to 300°C), free and substituted N-heterocyclics such as pyrroles, pyrrolidines, pyridines, pyranes, and pyrazoles, have been identified by analytical pyrolysis (Schulten et al 1997b). c) Their presence in humic substances and soils was also detected without pyrolysis by gel chromatography – GC/MS after reductive acetylation (Schnitzer and Spiteller 1986), by X-ray photoelectron spectroscopy (Patience et al. 1992), and also by spectroscopic, chromatographic, chemical, and isotopic methods (Ikan et al. 1992). 5. While we can see light at the end of the tunnel as far as soil-N is concerned, further research is needed to identify additional N-containing compounds such as N- heterocyclics, to determine whether these are present in the soil or humic materials in the form in which they were identified or whether they originate from more complex structures. If the latter is correct, then we need to isolate these complex N-molecules and attempt to identify them.     

GC-FID- and acyl carbon number-based determination of characteristic groupings of complex triglyceride (Benefat S and other) mixtures

Techniques for the gas and liquid chromatographic separation of complex mixtures of triglycerides have evolved over the past two decades, as reviewed in detail by Huang et al. (J. Agric. Food Chem. 1995, 43, 1834-1844; J. Agric. Food Chem. 1997, 45, 1770-1778). A novel method for the quantitative partitioning of complex mixtures of triglycerides into functionally related groups is developed and applied to a low-calorie triglyceride mixture [namely, Benefat S or Salatrim plus mid-chain (C(6,8,10,12)) fatty acids]. The method is based on a nonlinear calibration of retention times (RTs) of a suite of standard triglycerides on their acyl carbon numbers [(ACNs), the sum of all the acyl carbon atoms in a given triglyceride] to estimate all of the intermediate ACNs (from 6 to 66). With the calibrated ACN scale and identifications of some components of a complex mixture's composition, ACN-based partitions were established and a Benefat S-triglyceride chromatogram was partitioned into seven functionally related groups. This method is provisional in the sense that it would typically be employed when the identifications of many components of a complex, homologous series were unknown, yet functionally related groups needed to be quantified. This method has proven to be particularly useful in the intercalibration of research laboratories with production facility laboratories during complex ( approximately 50-90 compounds) and large-scale ( approximately 20 ton) syntheses because of the high reproducibility of the ACN-based partitioning of complex chromatograms. This carbon number and statistically based method can be generally applicable to other complex mixtures of organic compounds and is readily adaptable to laboratory intercalibration efforts.     

The involvement of iron and aluminum in the bonding of phosphorus to soil humic acid

Abstract

With a peat soil similar amounts of phosphorus (P) were coprecipitated with humic acid from alkali extracts over a limited range of strongly acidic pH, whereas with a mineral soil the amount was pH dependent. The difference between the two soils relates to the much greater total amounts of inorganic P and aluminum (Al) present in the extract of the mineral soil. In this acid mineral soil, Al rather than iron (Fe) may be involved in the formation of metal bridges in humic acid‐metal‐inorganic P complexes. Neither Al or Fe were implicated in binding of organic P to humic acid. The P species observed in humic acids was dependent on the pH at which they were precipitated from the alkali extracts. In the peat soil the inorganic P was an order of magnitude lower than the organic P.     

On artificial humic acids

I. Absorption Spectra of Some Artificial Humic Acids

It is a well known fact that numerous dark brown or black and amorphous substances cap be produced from organic compounds by purely chemical processes. Some of them are called artificial humic acids, and have been used as a model of soil humic acids by many workers. But it is doubtful whether such artificial humic acids are considered to be a model of soil humic acids, even if there can be found any similarities in their properties. However it may be an useful method for the study of humus formation to compare artificial humic acids with soil humic acids, and to make efforts to find a good model or to prepare artificial humic acids comparable to soil humic acids under laboratory conditions. For this purpose, first of all, the author prepared artificial humic acids from glucose, hydroquinone and lignin, and their absorption spectra were determined.     

Effect of aluminum on callose synthesis in root tips of tea (Camellia sinensis L.) plants

Aluminum (Al) toxicity is a major factor limiting yield production on acid soils (Foy 1983). The initial symptom of Al toxicity in many plants is manifested by the inhibition of root elongation (Ownby and Popham 1990; Llugany et al. 1994; Sasaki et al. 1994; Horst et al. 1997), which occurs during a very short period of time after exposure to Al (Llugany et al. 1994; Staß and Horst 1995). In a large number of recent reports, it was shown that the root apex plays a major role in the Al-sensitivity and response mechanisms (Zhang et al. 1994; Sasaki et al. 1997; Sivaguru and Horst 1998). However, it is interesting to note that stimulatory effects of Al on the growth of plants have also been reported in some studies (Chenery 1955; Konishi et al. 1985; Huang and Bachelard 1993; Osaki et al. 1997). In tea plant (Camellia sinensis L.) a stimulatory effect of Al on the growth was also demonstrated in some experiments, using intact plant (Chenery 1955; Konishi et al. 1985), cultured roots (Tsuji et al. 1994), and pollen tubes (Yokota et al. 1997). The growth of tea roots was typically more stimulated than that of shoots by Al (Konishi et al. 1985). It was assumed that Al effects might be due to the amelioration of phosphorus absorption (Konishi et al. 1985), secretion of malic acid from roots to dissolve aluminum phosphate in the rhizosphere (Jayman and Sivasubramaniam 1975), stimulation of growth of microorganisms on the root surface (Konishi 1990) or replacement of some functions of boron (Konishi 1992; Yokota et al. 1997). However, the stimulatory effects of Al on tea plant growth have not yet been el ucidated.

The formation of callose (1,3-β-glucan) has been reported as a common plant response to a variety of stresses, as well as mechanical, biophysical, chemical, and biological injury (Jaffe and Leopold 1984; Zhang et al. 1994). Increased synthesis of callose has been observed upon exposure to excess amounts of some elements, such as boron (McNairn and Currier 1965), cobalt, nickel, zinc (Peterson and Rauser 1979), and manganese (Wissemeier and Horst} 1987, 1992). Callose synthesis was also induced by Al in the roots of Triticum aestivum (Zhang et al. 1994) and Zea mays (Horst et al. 1997; Sivaguru and Horst 1998), suspension-cultured cells of Glycine max (Staß and Horst 1995), and protoplasts of Avena sativa (Schaeffer and Walton 1990) and Zea mays (Wagatsuma et al. 1995). Induction of callose synthesis in roots seems to be a very rapid physiological indicator of Al-induced injury or genotypical differences in Al sensitivity (Wissemeier and Horst 1992; Zhang et al. 1994; Horst et al. 1997). Nevertheless, Al-induced callose synthesis in tea plant, whose growth is stimulated by suitable Al concentrations, has not been described yet. Therefore, to elucidate the physiological basic effects of Al on tea plant, callose synthesis affected by Al in the root tips of intact plants was analyzed in the present study.     

Elemental,functional infrared and free radical characterization of humic acid-type fungal polymers (melanins)

Summary Humic acid-type polymers (melanins) synthesized in culture media by the fungi Aspergillus glaucus, Eurotium echinulatum, Hendersonula toruloidea, Stachybotrys atra and Aspergillus sydowi were analysed for elemental composition, functional group content, infrared (IR) and electron spin resonance (ESR) properties. Results were discussed in comparison with range values referred for soil humic acids. The fungal polymers showed significant differences in carboxyl and nitrogen content and C/H atomic ratios, reflecting a different degree of condensation (aromaticity) among the various samples. IR analysis gave evidence of: (a) the predominant aromatic character of melanins from A. glaucus, E. echinulatum and H. toruloidea; (b) the high content of aliphatic and olefinic components of S. atra melanin; (c) the typical presence of amide bonds in the nitrogen-richest melanins from A. sydowi and H. toruloidea; and (d) the generally low amount of free carboxyl groups, which often appeared involved in hydrogen bonds. ESR spectra showed that all the melanins studied contained appreciable concentrations of organic free radicals of prevailing semiquinonic nature and of the same order of magnitude commonly measured in humic acids from soil and other sources. The free electron concentration was shown to be directly related to the C/H atomic ratio and to the degree of aromaticity shown by IR analysis. This indicated that the highest free radical content in the melanins from E. echinulatum and A. glaucus was associated with the highest presence of condensed aromatic structures. Humic acid-type polymers synthesized by soil fungi may, therefore, contribute to the total free radical content of soil humic substances and play important roles in all reactions involving free radicals in soils and related environments.     

Fractionation of humic acids according to their hydrophobicity, size, and charge-dependent mobility by the salting-out method

Humic acids (HAs) represent heterogeneous and polydisperse mixture of molecules that differ in their chemical structure, composition, and functional properties. Fractionation of HAs is of key importance for understanding their interactions with various organic and inorganic compounds, for studying their physiological activity, and for predicting their behavior in natural environments and agroecosystems. Existing fractionation methods are rather laborious and time consuming, which limits their application in fundamental science and industry. It is shown that fractionation of humic acids with ammonium sulfate ensures their preparative separation with respect to (a) hydrophobicity, (b) molecular size, and (c) charge dependent on the amount of functional groups. Salting out at the lowest and highest degrees of saturation with ammonium sulfate, upon which precipitation of the molecules occurs, makes it possible to separate humic acids into functionally different high-molecular-weight/hydrophobic and low-molecular-weight/hydrophilic fractions. The first fraction is characterized by a lower electrophoretic mobility than the second fraction. The weight percentage of the components coagulated at the lowest degree of salt saturation can be used as a quantitative parameter for comparing hydrophobic properties of humic acids. Salting out is recommended as a fast, simple, and cheap alternative to chromatographic methods for preparative separation of humic acids if large amounts of functionally different fractions need to be obtained.     

Further investigations on the chemistry of fungal “humic acids”

In view of the considerable interest in laboratory-prepared fungal “humic acids” as possible precursors or incorporated structural components of soil humic substances, we degraded four fungal “humic acids” by the relatively mild alkaline cupric oxide oxidation. The oxidation products were extracted into organic solvents, methylated, separated by thin-layer chromatography and identified on a gas chromatographic-mass spectrometric-computer system.Average yields of major degradation products were: (a) aliphatic compounds, 38 per cent; (b) benzene-carboxylic compounds, 25 per cent; and (c) phenolic compounds, 21 per cent. The remaining 16 per cent consisted of a number of dialkyl phthalates. Our data agree with those that we reported earlier when we degraded a number of fungal “humic acids” by the more drastic alkaline permanganate oxidation and show that fungal “humic acids” are enormously complex organic materials containing aliphatic and aromatic structures, (some of which contain N), but only a relatively small proportion of which is phenolic. Most of the aliphatics isolated consisted of alkanes and fatty acids, which are known to persist in soils over long periods of time and are frequently firmly retained by soil humic substances.     

Role of inorganic anions in the structural transformation of molecules of humic substances in soil

It was shown that inorganic anions of exogenic compounds affected the structure of humic and fulvic acids in gray forest soil, as well as the yield of humic acids and their contents of carbon and metals. Humic substances in arable soil more readily interacted with the anions of extractants and fertilizers compared to virgin soil. Changes were revealed in the energy state of the electronic system of P- and N-modified fragments of humic acids from the arable soil. New ligand groups and chelate nodes were found in modified fragments using differential electronic spectra.     

The absorption and translocation of amino acids and amides in rice plant

It has been well known that the inorganic nitrogen compounds used as the common nitrogen source for the growth of higher plants can be replaced by some organic nitrogen compounds such as amino acids or amides. According to GHOSH and BURRIS (1), who investigated the effect of some amino acids as the nitrogen source, alanine, asparagine, glutamate and histidine were better nitrogen sources than ammonia for clover and tomato plants. For tobacco, however, nitrate and ammonia were superior to all organic nitrogen compounds used. RATNER et al. (2). made the same kind of study by using corn and sunflower plants and reported that the plants could grow with glycine, aspartate, glutamate and arginine, but all of them were inferior to inorganic nitrogen as the nitogen source.     

Factors affecting soil organic matter turnover in a mediterranean ecosystems from sierra de gador (Spain): An analytical approach

Abstract

Humus formations from forest and brushwood ecosystems on calcareous substrates in representative sites from Sierra de Gador in Southern Spain were analyzed. The humus composition, as determinated by three independent fractionation procedures, suggests that soil organic matter turnover is not related to the vegetation biotypes, but to a series of local factors among which climatic and soil characteristics predominate. Of the parameters studied, the amount of fulvic acids, and to a lesser extent, the relative proportion of soil perylenequinonic pigments (P‐type humic acids), reflected differences in altitude. The percentage of nitrogen (N) in humic acids was the parameter that was most informative of the characteristics of humus. This finding contrasts with the poorly differentiated patterns of the particulate organic matter fractions, whose evolution is probably independent of environmental factors because of physical protection by soil carbonates. The carbon dioxide (CO₂) release curves reproduced under laboratory conditions showed that the intrinsic biodegradability of soil organic matter does not reflect the nature of the plant residues in the sites sampled (170–2240 m), but is weakly dependent on climatic factors. We postulate that the calcium saturation, together with the active participation of inorganic colloids, force the convergent evolution of soil humus by compensating most of the effects of the climatic and biotic factors, and the historic influence of human activity in the area studied.     

Evaluation of the possibilities of using humic acids obtained from lignite in the production of commercial fertilizers

Purpose

Technological progress and high market demand contributed to a significant interest in the production of fertilizers based on humic acids. The aim of this study was to evaluate the possibilities of using humic acids obtained from lignite in the production of new commercial products. For this purpose, it is necessary to determine the quality standard requirements for such material. Properties of humic acids depend on source of origin as well as method of its extraction.

Materials and methods

The humic acids were extracted from polish deposit of lignite–Sieniawa Lubuska by alkaline extraction using for this purpose six kinds of extractants: 0.1 M NaOH and 0.25 M NaOH, 0.1 M KOH and 0.25 M KOH, and 0.1 M Na₄P₂O₇ and 0.25 M Na₄P₂O₇. The humic samples were used in solid powder form and characterized by UV-Vis spectroscopy, ¹³C NMR spectroscopy, fluorescence spectroscopy, and thermal analysis.

Results and discussion

The determining factor influencing a degree of humic acids extraction from lignite and their structure is type of extractant. The largest efficiency of extraction (about 50%) was obtained with the use of NaOH solutions. All examined humic acids were generally characterized by simple and heterogeneous molecularly structure with low molecular weight and low aromatic polycondensation. Therefore, it can be concluded that humic acids extracted with NaOH and KOH solutions are less condensed than those extracted with Na₄P₂O₇ solutions. It can suggest that humic acids obtained from lignite using solutions of Na₄P₂O₇ are characterized by a low transformation degree and greater amount of carboxyl groups.

Conclusions

Low rank coal can be successfully used in agriculture as a rich source of humic acids. Reagent used in their extraction, apart from high efficiency should have a neutral impact on their structure. Studies on the physicochemical properties of humic acids can be helpful in predicting behaviors of such fertilizer components in the environment and in inventing new products taking the principles of sustainable development into consideration.