1. Introduction
Additions of organic manures result in increased soil organic matter content. Many reports have shown that this results in increased water holding capacity, porosity, infiltration capacity, hydraulic conductivity and water stable aggregation and decreased bulk density and surface crusting [
1]. A straw return is the main method of crop straw treatment. However, the straw return method commonly used has many adverse effects on the levels and improvement of soil fertility and crop yield [
2]. The application of mineral fertilizers also results in an increase in the amount of organic matter returned to the soil [
1].
There are two means to increase the organic matter content in soils; one is to increase the organic matter gains or additions to the soil, and the other is to decrease organic matter losses. Storage of soil organic carbon is a balance between carbon additions from non-harvested portions of crops and organic amendments and carbon losses, primarily through organic matter decomposition and release of respired CO
2 to the atmosphere. Organic matter returned to the soil, directly from crop residues or indirectly as manure, consists of many different organic compounds. Some of these are digested quickly by soil microorganisms. The result of this is a rapid formation of microbial compounds and body structures, which are important in holding particles together to provide soil structure and limit soil erosion, and the release of carbon dioxide back to the atmosphere through microbial respiration. Thus, soil organic matter is not only an important source of carbon for soil processes but also a sink for carbon sequestration. Cultivation can reduce soil organic carbon content and lead to soil deterioration, and finally reduce soil productivity [
3].
The favorable quality of the humus positively influences the stabilization of water-stable soil aggregates, which are the basic units of the soil structure. As a consequence of soil cultivation, macroaggregates are broken into microaggregates which are stabilized mostly by humic acid carbon with highly condensed and stabilized macromolecules [
4].
Humic substances are a major stable part of soil organic carbon that play a central role in soil carbon accumulation [
5]. Humification depends on soil organic matter contents [
6] and the C/N ratio of a particular fertilizer [
7]. The continuous application of farmyard manure to field soils generally results in a higher humic acid content compared to the application of mineral fertilizers alone [
8]. Furthermore, a regular application of rotted or composted farmyard manure within the rotation can increase soil organic carbon content much more than the separate application of straw and cattle slurry [
5].
Both humic and fulvic acids have high sorption capacity with respect to many contaminants, including heavy metals, which can result in their immobilization and consequently protection of food and groundwater against contamination [
9]. The increase in water-soluble sugars, “non-matured” lignin and fulvic acids is an indicator of a labile system characterized by a rapid course of changes and a longer period of stabilization or acquisition of dynamic equilibrium in the mineralization and humification process [
10]. Furthermore, the high humic acid to fulvic acid ratio may explain decreased concentrations of metals in plants [
11]. Heclik et al. [
12] describe this phenomenon in their study on nanoparticles; fulvic acid molecules only form a salt with heavy metal ions, while the conformation of humic acid molecules is responsible for metal ion capture.
The E4/E6 ratio is inversely related to the degree of condensation of the aromatic network in humic acids. A low E4/E6 ratio is indicative of a relatively high degree of aromatic constituent condensation while a high ratio reflects a low degree of aromatic condensation and the presence of relatively large proportions of aliphatic structures [
13]. Therefore, the humic to fulvic acid ratio is increased together with decreasing values of the E4/E6 ratio measured in the visible spectrum range [
14].
Reliable quantitative evaluation of humic substances formation using, for example, parameters such as the humification index, humification degree and humification rate requires data from long-term experiments which are lacking because they are usually costly and time-consuming [
9]. Therefore, this study aims to evaluate the long-term application of mineral fertilizers, farmyard manure and plant residue incorporation on the quality of soil organic matter.
2. Materials and Methods
Long-term on-farm trials have been established within the years 1993 and 2000 by the Central Institute for Supervising and Testing in Agriculture at ten experimental sites with various soil-climatic conditions in the Czech Republic. The characteristics of the experimental sites are given in
Table 1. The average total organic carbon content varied between 8.1 and 15.0 g/kg. Within these trials, six crops were rotated in the following order: pea, canola, winter wheat, spring barley (1), beet/potato, and spring barley (2).
As follows from
Table 2, six treatments were studied: unfertilized treatment (unfert.), mineral fertilization (NPK)–basal application of Ca(H
2PO
4)
2 + KCl + (NH
4)
2SO
4 and top-dressing of calcium ammonium nitrate, application of farmyard manure (FYM), a combination of farmyard manure and mineral fertilization (FYM + NPK), incorporation of plant residues (STRAW/BT) and a combination of plant residues incorporation and mineral fertilization (STRAW/BT + NPK). The supply of nutrients in mineral fertilizers respected both the demands of crops and the maintenance of the optimal content (‘good’) of the available nutrients (Mehlich 3) nutrients in soil (K 170–310 mg/kg, P 80–115 mg/kg). At the sites of Lednice, Pusté Jakartice and Věrovany, beet was grown instead of potato, which resulted in the incorporation of the beet tops into the soil (
Table 1 and
Table 2). The incorporation of cereal and canola straw was accompanied by the application of 40 kg N/ha and 20 kg N/ha, respectively. Each treatment had four replicates. Furthermore, each replicate was repeated three times during the soil analysis.
Due to missing data related to the composition of organic fertilizers applied in all years of the experiments, the following parameters in dry matter were used to calculate carbon input and the C/N ratio: cereal straw 44% C [
15,
16] and a C/N ratio of 82 [
15,
17], pea straw 45% C [
18,
19] and a C/N ratio of 25 [
20,
21], beet tops 37% C [
22,
23] and a C/N ratio of 16 [
21,
23], farmyard manure 35% C [
24,
25] and a C/N ratio of 30 [
26,
27].
Soil samples were collected in 2016 after spring barley harvest (1) and analyzed as follows: A sample of 5.0 g of soil was stirred for 10 min in a mixture of 0.1 mol/L sodium pyrophosphate and 0.1 mol/L sodium hydroxide solution. After 24 h of storage, a saturated solution of sodium sulfate was added. A filtration followed. The filtrate formed was used for:
The E4/E6 ratio measurement directly in the filtrate. For the E4/E6 ratio, a visible light spectrometer Lambda 25 (PerkinElmer, Waltham, MA, USA) was used to calculate the specific spectral absorbance ratio at 465 and 665 nm [
28].
Determination of carbon in humic substances. The filtrate was neutralized by sulphuric acid and then vaporized. Iodometric titration followed.
Determination of carbon in fulvic acids. The filtrate was acidified by sulphuric acid to a pH of 1.0–1.5 and warmed up for 30 min. After storage for 24 h, the solution was filtrated and washed using the 0.05 mol/L sulphuric acid solution. The newly formed filtrate was vaporized. Iodometric titration followed.
The carbon of humic acids was determined. The remaining precipitate was dissolved using the hot 0.05 mol/L sodium hydroxide solution. After neutralization by sulfuric acid and vaporization, iodometric titration followed.
Before titration of all samples, dry matter formed by vaporization was dissolved in a mixture of the 0.067 mol/L potassium dichromate solution and concentrated sulfuric acid and warmed up for 45 min.
Humification indices were calculated according to Raiesi [
29] and Iqbal et al. [
30]:
where C
FA is the fulvic acid carbon, C
HA is the humic acid carbon and TOC is the total organic carbon in soil. The total organic carbon content in soil was determined from about 50 mg of soil by the modified Dumas combustion method at 960 °C with a CHNS Vario MACRO cube analyzer (Elementar, Langenselbold, Germany).
Except for the humification index, the studied variables significantly differed among experimental sites. Therefore, the effect of treatments was also evaluated by replacing the current values of variables with relative ones. Relative values were calculated as:
where V
treatment was the value of each treatment, and V
site-average was the average value of a particular site among all treatments.
Relative contribution (RC) of organic fertilizers to soil organic carbon stock was calculated according to Wang et al. [
31]. The unfertilized treatment (0) and the NPK treatment, respectively, were taken to be the “control”; the FYM and STRAW/BT treatments were compared with the unfertilized treatment, whereas the FYM + NPK and STRAW/BT + NPK treatments were compared with the NPK treatment.
The carbon sequestration efficiency (CSE) was calculated as follows:
where TCI is the total C input (t/ha) applied in organic fertilizers during the duration of individual experiments [
31].
A one-way analysis of variance (ANOVA) using Fisher’s LSD test was calculated. Pearson’s correlation coefficients were used to analyze the relationships among the variables studied in Table 4. The probability value of 0.05 or less (p < 0.05) was considered statistically significant. A statistical analysis of the data was carried out using the Statistica version 13.3 software (TIBCO Software, Palo Alto, Santa Clara, CA, USA).
3. Results
As is shown in
Table 3, unlike the content of fulvic acid carbon (C
FA), the content of humic acid carbon (C
HA) correlated significantly with the weighted average of the C/N ratio of applied organic fertilizers, RC and CSE (moderate correlations).
The E4/E6 ratio correlated significantly with the humic to fulvic acid carbon (CHA/CFA) ratio, although this correlation was weak. Although the E4/E6 ratio was significantly correlated with carbon input in organic fertilizers (moderate correlation), the CHA/CFA ratio was significantly correlated with the C/N ratio of organic fertilizers (weak correlation). The long-term annual average of both precipitation amount and air temperature was moderately correlated with both the CHA/CFA and the E4/E6 ratio. Higher values of the CHA/CFA ratio and lower values of the E4/E6 ratio were recorded under conditions of lower precipitation amount and higher air temperature. A stronger correlation was found between the CHA/CFA ratio and the CHA content, rather than the CFA content.
Unlike the humification rate, the humification index correlated significantly with the weighted average of the C/N ratio of applied organic fertilizers (moderate correlation), RC (weak correlation) and CSE (weak correlation).
Both the relative contribution of organic fertilizers to soil organic carbon stock (RC) and carbon sequestration efficiency (CSE) correlated positively with the CHA content and negatively with both carbon input in organic fertilizers and the weighted average of the C/N ratio of applied organic fertilizers. All these correlations were moderate.
Carbon input did not correlate significantly with any fractions of soil organic carbon in soil. In contrast, the weighted average of the C/N ratio of applied organic fertilizers correlated significantly with the CHA content and the humification index (moderate correlation), and with the CHA/CFA ratio (weak correlation).
3.1. Organic Carbon Fractions
The relative values of studied variables independent of the site effect are often mentioned in the following tables (
Table 4,
Table 5 and
Table 6). The FYM + NPK treatment led to an increase in the relative C
FA content compared to the NPK and FYM treatments (
Table 4). Except for the FYM + NPK treatment, no significant differences in the C
FA content among other treatments were found.
The FYM + NPK treatment resulted in an increased relative CHA content compared to the unfertilized treatment, NPK treatment, STRAW/BT and STRAW/BT + NPK treatment. The STRAW/BT + NPK treatment led to a decrease in relative CHA content compared to all other treatments except the unfertilized. Even though the relative CHA content did not differ between the FYM treatment and the STRAW/BT treatment, in absolute figures, the FYM treatment achieved higher CHA content compared to the STRAW/BT treatment at half of the experimental sites.
Compared to the unfertilized treatment, the relative C
HA/C
FA ratio was increased in the NPK, FYM and FYM + NPK treatments (
Table 5). The FYM + NPK treatment resulted in an increased relative C
HA/C
FA ratio in comparison with the STRAW/BT + NPK treatment. A decrease in the relative C
HA/C
FA ratio was recorded in the STRAW/BT + NPK treatment compared to the NPK treatment. Even though the relative C
HA/C
FA ratio did not differ between the FYM and the STRAW/BT treatment, in absolute figures, a higher C
HA/C
FA ratio in the FYM treatment compared to the STRAW/BT treatment was recorded at four experimental sites.
The highest E4/E6 ratio was found in the FYM treatment at half of the experimental sites. Lower values of the relative E4/E6 ratio were recorded in the STRAW/BT treatment in comparison with the FYM treatment. However, no significant difference in relative E4/E6 ratio between the FYM + NPK and STRAW/BT + NPK treatment was recorded.
3.2. Degree of Humification
The FYM + NPK treatment resulted in an increased relative humification rate compared to all other treatments except for the unfertilized one (
Table 6).
In terms of the relative humification index, the FYM + NPK treatment achieved higher values in comparison with all other treatments while the STRAW/BT + NPK treatment resulted in a lower relative humification index compared to all treatments except for the STRAW/BT one. Even though no significant difference in relative humification index between the FYM and the STRAW/BT treatment was found, in absolute numbers, a higher humification index in the FYM treatment compared to the STRAW/BT treatment was recorded at half of the experimental sites.
3.3. Carbon Sequestration
The influence of treatment was recorded on neither the relative contribution of organic fertilizers to soil organic carbon stock (RC) nor carbon sequestration efficiency (CSE) at only two experimental sites (
Table 7). Unlike the straw return, farmyard manure application resulted in higher (positive) values of both RC and CSE. On average, 30.75% and 43.20% of the carbon input in farmyard manure was converted to the organic carbon content of the soil in the FYM and the FYM + NPK treatment, respectively.