Chemical and physical characteristics of humic substances extracted from greek soils varying in classification order and natural drainage

Abstract

Humic and fulvic acids were extracted from the surface horizons of Alfisols, Vertisols, and Entisols with good and impeded natural drainage. These profiles are located in the temperate subhumid zone. The extracted humic substances were characterized by elementary and functional group analyses, IR‐spectroscopy, differential thermal analysis (DTA) and electron spin resonance spectroscopy (ESR). Some differences in the humic substances seem to be associated with differences in the natural drainage of the soils. When data from samples of the same soil order are compared, indicate that humic (HAs) and fulvic acids (FAs) extracted from soils with impeded drainage, have higher contents of carbon and spin concentration, but less total acidity and fewer carboxylic groups (COOH), than the respective humic substances extracted from well drained soils. The IR‐spectra of the HAs extracted from soils with impeded drainage showed more aliphatic groups than those of the well drained soils. No essential differences, which could be related to the drainage conditions of the studied soils, were observed in the IR‐spectra of FAs. DTA‐diagrams shows that the thermal stability of the HAs and the FAs are related to their carbon and ash contents. Iron was the dominant element in the ash of the humic acids, aluminum and silicon were the most abundant elements in the ash of the fulvic acids.     

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.

     

Chemical studies on soil humic acids

Seventeen samples of soil humic acids, two fractions of soil fulvic acid sample, and several related compounds such as lignin, tannin, flavonoid and artificial humic substances were decomposed in conc. KOH solution at 180°C. Succinic acid, glutaric acid, phloroglucin, p-hydroxybenzoic acid, vanillic acid, protocatechuic acid, 3,4-dihydroxy-5-methoxybenzoic acid, and gallic acid were detected in the degradation products of humic acids. The amounts of these degradation products were discussed in relation to the degree of humification or the sources of the humic acid samples. Succinic acid also resulted from glucose, polymaleic acid, and the humic acid and humin prepared from glucose, but glutaric acid resulted only from glucose humic acid and glucose humin but not from glucose and polymaleic acid. Succinic acid and glutaric acid were supposed to result from the same structural portions in humic acids because of the very significant positive linear correlation between their amounts. p-Hydroxybenzoic, vanillic, protocatechuic, and 3,4-dihydroxy-S-methoxybenzoic acids were presumed to result mainly from lignin structure in humic acids. Soil humic acids yielded small amounts of gallic acid although the yields by hydrolysable tannins were in large amounts. The yields of above-mentioned degradation products from humic acids decreased with increasing degree of humification. Phloroglucin resulting from ftavonoids including condensed tannins were also found in the degradation products of humic substances. Its yield showed no linear correlation with RF value of humic acid, and is presumed to be rather related to the vegetation at the sites of soil sampling.     

A model for a type humic acid

In the preceding papers (1,2,3,4), the author studied some chemical properties of soil humic acids, and compared various so-called artificial humic acids with soil humic acids in respect to their absorption spectra and stabilities. According to these experimental results, it is Presumed that soil humic acids, especially A type, must contain not only benzene ring, but also condensed aromatic rings as their structural units.     

Temporal changes in humic acids in cultivated soils with continuous manure application

Abstract

Although the application of manure to upland fields is believed to induce changes in the quality of humic substances in soil as well as the quantity, the direction and extent of these changes have not been elucidated. To understand temporal variations in humic acids, periodically collected soil samples from two fields, a Typic Hapludult (Togo) and a Pachic Melanudand (Kuriyagawa), with cattle manure and chemical fertilizer (CF) were examined. The content and degree of humification (darkening) of the humic acids were distinctly greater in Kuriyagawa than in Togo soil. Corresponding to the difference in the degree of humification, molecular size distribution, elemental composition, infrared (IR) spectra, and ¹³C cross polarization/magic angle spinning nuclear magnetic resonance (CPMAS NMR) spectra of humic acids differed between the two soils. Manure application at 40 Mg ha^?1 year^?1 for 16 years (Togo) and at 80 or 160 Mg ha^?1 year^?1 for 19 years (Kuriyagawa) resulted in greater humic acid content compared with plots with CF only because of its increase in the manured plots and/or decrease in the CF plots. Manure application at an extremely high rate (160 Mg ha^?1 year^?1) resulted in higher H content and greater signal intensities of alkyl C, O-alkyl C and amide C=O in the ¹³C CPMAS NMR and/or IR spectra. Although humic acids with larger molecule sizes increased in all the manured plots, differences between the humic acids from the plots with and without manure applied at practical levels in the elemental and spectroscopic analyses were small or scarce. These results were considered to be because of the similarity between the indigenous soil humic acids and the manure-derived ones in Togo soil (a low degree of humification) and because of the abundance of highly-humified humic acids in Kuriyagawa soil.     

Short-term impact of spent coffee grounds over soil organic matter composition and stability in two contrasted Mediterranean agricultural soils

Purpose

Spent coffee grounds (SCG) is a biowaste which arouse great interest as soil organic amendment due to the huge amount produced around the world. However, the impact of this residue on soil organic matter (SOM) functionality and stability has been barely studied. Thus, the aim of this work is to study the short-term effects of SCG on the quantity and quality of SOM in two Mediterranean agricultural soils (Vega soil, SV and Red soil, SR) in microcosm conditions.

Materials and methods

The in vitro assay was performed with two fresh SCG doses (2.5 and 10% w/w), two incubation times (30 and 60 days) and two agricultural soils (SV and SR). SOM fractionation to obtain total extractable carbon, humic acids, fulvic acids, humins and hot water soluble carbon (HWSC) was determined. Spectroscopic UV-Vis and Mid-IR, thermogravimetric and simultaneous differential thermal analysis, as well as scanning electron microscopy (SEM), were also applied in this study.

Results and discussion

SCG increased all SOM fractions, especially the levels of more labile SOM (HWSC, increased 600–700%) and total extractable carbon (increased to around 200%). SCG also increased humic acids and fulvic acids around 200%, but the functionality of humic acids was affected by a reduction of the functional groups with more recalcitrant and stable character. The tested soils are different from each other (the SV has a more clayish texture and a higher smectite clay content than the SR) which made the behaviour of these soils different. The degree of incorporation of SCG into the soils structure and the interaction between soil and SCG particles (observed by SEM) affected carbon retention under stable forms, increasing carbon stabilization in SV with respect to SR.

Conclusions

The short-term effect of SCG on SOM composition and functionality demonstrate that this bioresidue could be used as soil organic amendment, being a valuable alternative use of a polluting waste. Soil type is a key factor since it influences the soil-SCG interaction and consequently SOM stability. To deepen the study of those effects, it would be necessary to analyze the long-term effects, field studies and to test in a greater number of soil types.

     

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.     

Effect of Application of Sulfuric and Humic Acid in the Mitigation of Root Asphyxiation Stress in Potted Avocado Plants

ABSTRACT

Avocado crop is very sensitive to root asphyxiation. Among the agricultural management techniques, soil applications of humic and sulfuric acids became increasingly popular. In a trial with potted 'Hass' avocado trees grown on a silty-clay soil, different soil treatments were applied to determine their effects on tree physiology and soil characteristics: Control, deionized water; SA, H₂SO₄ application; HA, humic acid application; SA+HA, combination of both treatments. Treatments were applied during a 9-month period. SA reduced soil pH, rised electrical conductivity and reduced saturated soil hydraulic conductivity (Ks) in the upper layers of the potted soil. On the other hand, HA increased electrical conductivity and Ks compared with Control. None of the treatments improved CO₂ assimilation, stomatal conductance, stem water potential, chlorophyll fluorescence, growth or nutrient content. Our findings suggest that a rapid and positive response to the application of acids is unlikely to be observed under conditions of restricted oxygen in the root zone.     

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.     

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.     

Formation of humic acids during plant residue humification and their nature

The process of humic acid formation during humification of plant residues (clover leaves and roots as well as timothy roots) was studied. According to some characteristics (elementary composition, IR-spectra, the content of COOH-groups) the newly-formed humic acids may be defined as “young forms”. The comparison of the new forms with more mature soil humic acids revealed the following specific features of the latter: (a) a higher degree of aromatic nuclei condensation; (b) a higher content of carboxyl groups; and (c) a more uniform structure.The authors discuss some defects of the methods used for extraction of humic acids from fresh and slightly humified plant residues by alkaline solutions. An artificial formation of humus-like substances and their extraction as admixtures together with true humic acids are inevitable with these methods. This should be taken into account in studying and systematizing such forms of humic acids.     

Quantitative and qualitative characterisation of humic products with spectral parameters

Purpose

The application of different humic products for the treatment of soils and plants has increased in recent years. The characteristics of humic products, such as the content and composition of organic carbon and the maturity, provide valuable information which is essential for an adequate application. Such information is crucial for manufacturers, business consultants and users involved in the production, distribution and implementation of humic products. This article presents the correlation between the quantitative indicators of commercial humic products and their spectral characteristics via measurements in the ultraviolet spectrum at 300 nm, in the visible area at 445 and 665 nm and in the near-infrared spectrum at 850 nm.

Materials and methods

We evaluated humic products (liquid and solid) of different origins. Via wet combustion, the content of total organic carbon in humic products can be determined. The precipitation of humic acids from the starting solution determines the composition of the humic products in terms of humic acids (HAs) and fulvic acids (FAs). The dissolution of HAs determines their concentration by titration, while the specific extinction can be assessed via spectrophotometry via measuring the absorption of HAs spectra at the following wavelengths: 300, 465, 665 and 850 nm. The degree of aromaticity and condensation of humic products determines the optical density of the HAs via the E₄/E₆ ratio.

Results and discussion

The content of total organic carbon varied widely from 0.55 to 37.5% across all groups. The content of carbon in HAs, as a percentage of the total carbon in fulvic-type humic products, ranged from 1.29 to 16.00%, while in humic-type products, it ranged from 51.43 to 91.92%. The minimum value of the E₄/E₆ ratio was 2.97, while the maximum value was 6.35. We observed a direct relationship between the dominant type of acids in humic products and the E₄/E₆ ratio.

Conclusions

The optical density of HAs indicates their quality characteristics. The presented optical characteristics for humic products show that there is a direct relationship, especially between HAs/FAs and E₄/E₆ ratios. Measurement at 300 nm (E₃₀₀) in the near-ultraviolet area and at 850 nm (E₈₅₀) in the near-infrared area can increase the range of the spectral study.

     

Effects of humic acid fractions on invertase activities in plant tissues

The effect of some soil organic matter fractions and synthetic humic acids were tested on the invertase activities in tissues of several higher-plant species. The effects varied according to the plant and soil organic matter fraction used. Thus no fraction tested had any effect on the invertase activity of beetroot storage tissue, but several fractions, mainly humic acid and its residues obtained by water or acid boiling, inhibited invertase in carrot storage tissues and wheat roots and coleoptiles. and to a lesser extent mungbean hypocotyls. Soil phenolic acids were without effect in any of the plant tissues tested. Some stimulations of invertase activity occurred with pea root tissue, using the water- and acid-soluble humic-acid fractions. The effects were independent of ash contents, total N or the C:H:N ratio, and could not be ascribed to any particular chemical component.The inhibitory effects on wheat invertase activity produced by soil organic matter were independent of pH in the range 4.5–7.0, and were constant over a 4 h incubation. The inhibitors only slightly affected the K_m but markedly affected the V_max of the reaction, hence they were non-competitive.The effect of a naturally-occurring invertase inhibitor from beetroot storage tissues could be reduced in the presence of humic acid. Although the inhibitor had little effect on wheat root invertase it did reduce the inhibitory effect of the humic acid in this tissue.     

Abbau von Huminstoffen durch Bodenmikroorganismen — eine Übersicht

Degradation of humic substances by soil microorganisms — a review Humic substances which represent differently extractable fractions of the soil organic matter exert multifarious effects on soil as a site for plant growth and a part of terrestrial environments. Among them especially humic acids and fulvic acids are subject to degradation and/or transformation by soil microorganisms. Several authors demonstrated the participation of different species of fungi, actinomycetes and also of non-mycelial aerobic or anaerobic bacteria in those processes under laboratory conditions. Indications exist that humic substances irrespective of their structure undergo degradation on cell surfaces due to the activity of exoenzymes. In this respect microbial phenoloxidases play an extraordinarily important role. The degradation rate of humic substances can be followed by optical, gravimetric and chemoanalytical methods as well as using biochemical and microbiological procedures (CO₂ release, microbial growth, biomass formation). An objective evaluation, however, can be hindered by the adsorption of humic substances on microbial biomass and sometimes also by formation of novel humic-like microbial metabolites. Therefore it is necessary to apply a multifactorial approach in the study of the degradation of humic substances which includes both quantitative and qualitative parameters. To better elucidate how these processes may occur under natural conditions, mixed populations of soil microorganisms should be predominantly involved in future studies.     

Use of alkaline soil extracts for 13C n.m.r. characterization of humic substances

Previous solution ¹³C n.m.r. studies of soil organic matter have been confined to isolated humic or fulvic acid fractions. A study of 10 crude alkaline extracts from five New Zealand topsoils has now shown that such purification is unnecessary. Spectra were not noticeably broadened by co-extracted iron in concentrations up to 2 mg cm^?3. Optimized pulse parameters were similar to those reported for a solution of a purified humic acid. Relaxation results support a generalized model for humic material, in which aromatic structures, carbohydrates, amino acids and polymethylene chains are linked together to form flexible macromolecules. Relative contributions from each component varied from soil to soil.     

Macromolecule‐P associations and inositol phosphates in some chilean volcanic soils of temperate regions

Abstract

Total organic P, humic and fulvic acid‐P associations and inositol phosphates in nine volcanic soils of southern Chile were determined. The concentration of organic P (Po) ranged from 654 to 1942 ppm accounting for 49% to 64% of total soil P. Phosphorus associated to humic (HA‐P) and fulvic acids (FA‐P) accounted for 51–68% and 32–49% of Po, respectively. Inositol penta‐ and hexaphosphates represented 42% to 67% of Po suggesting that significant amounts are associated with both humic and fulvic acids. Po content was significantly correlated to organic C, total soil P and HA‐P. HA‐P and FA‐P fractions obtained from the most representative soil were examined by dyalisis and gel filtration. While approximately 96% of HA‐P presented a molecular weight higher than 100,000 daltons, 53% of FA‐P had a molecular weight under 12,000 daltons. It is suggested that these more labile organic P forms would be more easily mineralized, thus increasing the available P pool.     

Effects of humic and fulvic acids on release of fixed potassium

Release of potassium fixed by expanding silicate clays is considered of practical importance in soil fertility. Humic and fulvic acids are expected to play a definite role in liberating this fixed K, because of their chelating power, but not much is known in this respect. The following investigation was conducted to study release of fixed K by montmorillonite and illite, using humic and fulvic acids isolated from the surface horizon of a Cecil soil (Typic Hapludult, Red Yellow Podzolic soil) as extractants. For comparison, extraction was also done with 1 N neutral NH₄ -acetate, a mixture of 0.05 N HCl and 0.025 N H₂SO₄ or H₂ O. Supporting analyses of clays were carried out with X-ray diffraction to establish changes, if any, in crystal structure due to fixation, release of K, or adsorption of humic compounds. The results indicated that humic and fulvic acids released some of the K fixed by montmorillonite or illite. In terms of percentage of the total K fixed, 9 to 28% were released by the various extractants. The percentages K released by humic and fulvic acids were similar from both montmorillonite and illite, but based on absolute values, humic and fulvic acids extracted less K (mg/100 g) from illite than montmorillonite. Although statistically significant at the 5% level of probability, the capacity of humic compounds to liberate fixed K was not different markedly from those of NH₄ -acetate and the double acid mixture. Differences in pH of humic solutions had no influence on extraction of fixed K. X-ray diffraction analysis yielded curves showing an increase in spacing from 10.4 Å for K-montmorillonite to 13.2 Å as a result of extraction with the double acid mixture.     

Selective digestion of the proteinaceous component of humic substances by the geophagous earthworms Metaphire guillelmi and Amynthas corrugatus

Humic substances play a key role in the global carbon cycling and the sequestration of micropollutants in soil. The transformation of these substances by earthworms, the dominant soil macroinvertebrates of many terrestrial ecosystems, and the mechanisms involved are still obscure. We prepared two chemically identical humic model compounds that were specifically ¹⁴C-labeled either in the aromatic or the proteinaceous component, and added them to soil incubated with the geophagous earthworm species Metaphire guillelmi (anecic) and Amynthas corrugatus (endogeic). In the absence of the earthworms, both the aromatic and the proteinaceous components were mineralized at similarly low rates (5−8% after 9 days of incubation). In the presence of the earthworms, mineralization rate of the proteinaceous component was strongly stimulated (2-fold by M. guillelmi and 1.4-fold by A. corrugatus). The mineralization rate of the aromatic component was (slightly) stimulated (1.2-fold; P < 0.05) only by A. corrugatus. In all cases, the stimulated mineralization was accompanied by a transformation of radiolabeled humic acids to fulvic acids within the earthworm guts and by an incorporation of radiolabel into the earthworm tissues. Digestion of the proteinaceous component of humic acids by the earthworms was corroborated also by a decrease of extractable humic acids in fresh cast and a stimulated mineralization of soil nitrogen; in the case of M. guillelmi, the fresh cast contained sixfold more NH₄⁺ than the non-ingested soil. Our study provides direct evidence for the selective digestion of humic components by earthworms. Considering the ubiquity of geophagous earthworms and their large biomass, the alteration of the chemical structure of humic substances by the earthworms through their selective digestion of peptidic components may have significant impacts on the stability of humic substances and the bioavailability of micropollutants in soil.