**4. Impact of application of organic fertilizers on the soil properties**

It is well known that organic fertilizer amendments can enhance soil fertility, increase soil aggregation, and increase soil pH (e.g., acidic soils). [21–23] For example, long-term (over 20 years) organic fertilization or organic plus inorganic fertilization could markedly improve soil organic carbon (SOC) content when compared to no fertilization (Control) and chemical fertilization (NPK) (**Figure 10**). Using Nano-CT, the structure of long-term fertilized soils at the Jinxian Experiment station was examined (**Table 5**). It was found that long-term organic amendments improved soil aggregation by decreasing the number of pores, pore throats, and paths between adjacent nodal pores in soil aggregates (**Table 5**). [24]

**Indices (***x***) Regression equation Maturity value in**

OM *y* = 3.3 + 1.1 × 10−2*x* 30% 3.63 GI *y* = 4.0 − 6.8 × 10−3*x* 50% 3.66 OUR *y* = 3.7 + 3.6 × 10−3*x* 10 mg O2/g-OM/d 3.74 CER *y* = 3.6 + 8.9 × 10−3*x* 4 mg C-CO2/g-OM/d 3.64

OM *y* = 2.9 + 1.6 × 10−2*x* 30% 3.38 GI *y* = 3.8 − 6.8 × 10−3*x* 50% 3.46 OUR *y* = 3.5 + 3.9 × 10−3*x* 10 mg O2/g-OM/d 3.54 CER *y* = 3.4 + 8.8 × 10−3*x* 4 mg C-CO2/g-OM/d 3.44 TOC, total organic matter; OM, organic matter; GI, germination index; OUR, oxygen uptake rate; CER, CO2 evolution

Some investigators have shown great interest in the assessment of compost maturity using fluorescence intensity. Our results demonstrate that log scores of components 1 and 3, identified by the DOMFluor–PARAFAC approach, can be applied to assess the maturity of composts which cover a large range of waste sources. This is attributable to the fact that the full fluorescence spectroscopy analysis provides a basis for capturing subtle changes in the fluorescence spectra. In summary, the assessment of compost maturity by the DOMFluor–

**4. Impact of application of organic fertilizers on the soil properties**

paths between adjacent nodal pores in soil aggregates (**Table 5**). [24]

It is well known that organic fertilizer amendments can enhance soil fertility, increase soil aggregation, and increase soil pH (e.g., acidic soils). [21–23] For example, long-term (over 20 years) organic fertilization or organic plus inorganic fertilization could markedly improve soil organic carbon (SOC) content when compared to no fertilization (Control) and chemical fertilization (NPK) (**Figure 10**). Using Nano-CT, the structure of long-term fertilized soils at the Jinxian Experiment station was examined (**Table 5**). It was found that long-term organic amendments improved soil aggregation by decreasing the number of pores, pore throats, and

Component 1 (*y*) TOC *y* = 3.7 + 7.5 × 10−3*x* 10 mg/g-DM 3.78

12 Organic Fertilizers - From Basic Concepts to Applied Outcomes

Component 3 (*y*) TOC *y* = 3.5 + 1.2 × 10−2*x* 10 mg/g-DM 3.62

**Table 4.** Calculated log (scores) of components from regression equations (*n* = 60) [1]

PARAFAC approach is not wastesource–specific.

rate.

**the literature [9]**

**Calculated log (scores) of components**

**Figure 10.** Dynamics of soil organic carbon (SOC) with fertilization time at the Qiyang long-term fertilization experi‐ ment station.


Values in parentheses represent the standard deviation of the mean. Different letters following values between different fertilization treatments indicate significant differences at *P* < 0.05 (LSD). The soil at the Jinxian Experiment station is red soil.

**Table 5.** Summary of soil microstructure properties from three contrasting fertilization treatments at the Jinxian Experiment station, Jiangxi Province, China [24]

However, very few studies are conducted on how organic fertilizer amendments affect the morphology and coordinate state of nanominerals or organomineral associations in soil. Highresolution transmission electron microscopy (HRTEM) coupled to selected area electron diffraction (SAED) technique could be a promising tool to observe the morphology/appearance of nanominerals in soil dissolved organic matter (DOM). Under three fertilization treatments at the Qiyang Experiment station, HRTEM observation showed two regions (i.e., black and gray regions) of soil DOM with distinctive percentage presented under Control (no fertiliza‐ tion), NPK (chemical nitrogen, phosphorus, and potassium fertilization), and NPKM (NPK plus manure fertilization) at the 22 years' long-term fertilization site (**Figures 11**–**13**). The electron diffraction patterns indicated that the obtained nanominerals in the black and gray regions had a determinate crystalline and amorphous pattern, respectively. Elemental maps further confirmed that crystalline nanominerals were dominated by Fe and O, while amor‐ phous nanominerals were mainly composed of Al, Si, and O (**Figures 11**–**13**). These results showed that after 22 years' fertilization, crystalline nanominerals were predominant under NPK, while amorphous nanominerals under both the Control and NPKM.

**Figure 11.** High-resolution transmission electron microscopy (HRTEM) images of soil dissolved organic matter from Control fertilization test in the long-term (22 years) location experiment. [4] (a) TEM image; (b) HRTEM images and selected area electron diffraction (SAED) pattern of the two regions indicated by blue squares, indicating that the black region is completely crystalline, while the gray region remains amorphous; (c) elemental maps; (d) EDS image. Con‐ trol, no fertilization.

However, very few studies are conducted on how organic fertilizer amendments affect the morphology and coordinate state of nanominerals or organomineral associations in soil. Highresolution transmission electron microscopy (HRTEM) coupled to selected area electron diffraction (SAED) technique could be a promising tool to observe the morphology/appearance of nanominerals in soil dissolved organic matter (DOM). Under three fertilization treatments at the Qiyang Experiment station, HRTEM observation showed two regions (i.e., black and gray regions) of soil DOM with distinctive percentage presented under Control (no fertiliza‐ tion), NPK (chemical nitrogen, phosphorus, and potassium fertilization), and NPKM (NPK plus manure fertilization) at the 22 years' long-term fertilization site (**Figures 11**–**13**). The electron diffraction patterns indicated that the obtained nanominerals in the black and gray regions had a determinate crystalline and amorphous pattern, respectively. Elemental maps further confirmed that crystalline nanominerals were dominated by Fe and O, while amor‐ phous nanominerals were mainly composed of Al, Si, and O (**Figures 11**–**13**). These results showed that after 22 years' fertilization, crystalline nanominerals were predominant under

**Figure 11.** High-resolution transmission electron microscopy (HRTEM) images of soil dissolved organic matter from Control fertilization test in the long-term (22 years) location experiment. [4] (a) TEM image; (b) HRTEM images and selected area electron diffraction (SAED) pattern of the two regions indicated by blue squares, indicating that the black region is completely crystalline, while the gray region remains amorphous; (c) elemental maps; (d) EDS image. Con‐

trol, no fertilization.

NPK, while amorphous nanominerals under both the Control and NPKM.

14 Organic Fertilizers - From Basic Concepts to Applied Outcomes

**Figure 12.** High-resolution transmission electron microscopy (HRTEM) images of soil-dissolved organic matter from NPK fertilization test in the long-term (22 years) location experiment. [4] (a) TEM image; (b) HRTEM images and se‐ lected area electron diffraction (SAED) pattern of the two regions indicated by blue squares, indicating that the black region is completely crystalline, while the gray region remains amorphous; (c) elemental maps; (d) EDS image. NPK, chemical nitrogen, phosphorus, and potassium fertilization.

**Figure 13.** High-resolution transmission electron microscopy (HRTEM) images of soil-dissolved organic matter from NPKM fertilization test in the long-term (22 years) location experiment. [4] (a) TEM image; (b) HRTEM images and selected area electron diffraction (SAED) pattern of the two regions indicated by blue squares, indicating that the black region is completely crystalline, while the gray region remains amorphous; (c) elemental maps; (d) EDS image. NPKM, NPK plus manure fertilization.

To understand whether fertilization practices can affect the local coordination state and the environment of Al and Si, the 27Al and 29Si nuclear magnetic resonance spectroscopy (NMR) spectra of soil water-dispersible colloids were used (**Figure 14**). The results showed that octahedrally coordinated aluminum (VIAl), with a peak at approximately 0 ppm, was the only type of aluminum in the soil water-dispersible colloids under the 22-year long-term Control, M, and NPKM treatments (**Figure 14**). Although the octahedrally coordinated Al was also dominant in soil colloids under the NPK, NPKCa (NPK + lime), N, and NCa (N + lime), small amounts of distorted tetrahedrally coordinated aluminum (IVAl, 46 ppm) and pentahedrally coordinated aluminum (VAl, 25 ppm) were observed. Distorted IVAl and VAl are usually present in well-characterized crystalline minerals. [25–27] This finding strongly implies that chemical fertilization modified the local coordination state and environment of Al, with a small part of IVAl replaced by distorted IVAl and VAl, whereas organic fertilization or organic plus chemical fertilization did not influence the local coordination state and Al environment. The results from the high-resolution 27Al NMR spectra support the finding that amorphous Al is more present in organic fertilizations (i.e., M and NPKM) than in chemical fertilizations (i.e., NPK, NPKCa, N, and NCa) (**Figure 15**). 29Si NMR spectra also confirmed the presence of amorphous Al as allophane and imogolite in the soils under control, M, and NPKM, but not under the four chemical fertilizations (N, NCa, NPK, and NPKCa). These results from 27Al and 29Si NMR spectra are consistent with our previous publication, in which nanominerals were directly observed by HRTEM images of soil DOM. [4]

**Figure 14.** High-resolution 27Al and 29Si NMR spectra of water-dispersible colloids from the long-term fertilized soils. [28] Control, no fertilization; N, chemical nitrogen; NCa, chemical nitrogen plus lime; NPK, chemical nitrogen, phos‐ phorus, and potassium; NPKCa, chemical nitrogen, phosphorus, and potassium plus lime; NPKM, NPK plus swine manure; and M, swine manure. The results of 29Si NMR spectra support the presence of nanominerals in organic (i.e., NPKM and M) rather than chemical (i.e., N, NCa, NPK, and NPKCa) fertilization treatments.

Selective extraction also showed that the Al fractions were significantly (*P <* 0.05) altered by the long-term fertilization treatment (**Figure 10**). Significantly higher amorphous Al concen‐ trations among the fertilizers were ranked as M ≈ NPKM > NPK ≈ NPKCa > N > NCa > Control (**Figure 10**). Significantly higher strongly organically bound Al concentrations ranked as NPKM > Control > NCa > NPK > NPKCa > M > N; significantly higher weakly organically bound Al concentrations ranked as NCa ≈ NPKCa > NPKM ≈ NPK ≈ Control > M > N. In addition, significantly higher exchangeable Al concentrations ranked as N > NPK > NPKCa > NCa > Control > M ≈ NPKM. These four fractions followed the pattern: organically bound Al > amorphous Al fraction > exchangeable Al. The results demonstrated that organic fertilization treatments increased amorphous Al and reduced exchangeable Al compared with chemical fertilization treatments. The addition of lime significantly (*P* < 0.05) increased the weakly organically bound Al and reduced exchangeable Al, suggesting that lime amendment transferred Al fractions from exchangeable Al to the weakly organically bound Al. These trends definitely affected soil C sequestration and soil pH.

To understand whether fertilization practices can affect the local coordination state and the environment of Al and Si, the 27Al and 29Si nuclear magnetic resonance spectroscopy (NMR) spectra of soil water-dispersible colloids were used (**Figure 14**). The results showed that octahedrally coordinated aluminum (VIAl), with a peak at approximately 0 ppm, was the only type of aluminum in the soil water-dispersible colloids under the 22-year long-term Control, M, and NPKM treatments (**Figure 14**). Although the octahedrally coordinated Al was also dominant in soil colloids under the NPK, NPKCa (NPK + lime), N, and NCa (N + lime), small amounts of distorted tetrahedrally coordinated aluminum (IVAl, 46 ppm) and pentahedrally coordinated aluminum (VAl, 25 ppm) were observed. Distorted IVAl and VAl are usually present in well-characterized crystalline minerals. [25–27] This finding strongly implies that chemical fertilization modified the local coordination state and environment of Al, with a small part of IVAl replaced by distorted IVAl and VAl, whereas organic fertilization or organic plus chemical fertilization did not influence the local coordination state and Al environment. The results from the high-resolution 27Al NMR spectra support the finding that amorphous Al is more present in organic fertilizations (i.e., M and NPKM) than in chemical fertilizations (i.e., NPK, NPKCa, N, and NCa) (**Figure 15**). 29Si NMR spectra also confirmed the presence of amorphous Al as allophane and imogolite in the soils under control, M, and NPKM, but not under the four chemical fertilizations (N, NCa, NPK, and NPKCa). These results from 27Al and 29Si NMR spectra are consistent with our previous publication, in which nanominerals were directly

**Figure 14.** High-resolution 27Al and 29Si NMR spectra of water-dispersible colloids from the long-term fertilized soils. [28] Control, no fertilization; N, chemical nitrogen; NCa, chemical nitrogen plus lime; NPK, chemical nitrogen, phos‐ phorus, and potassium; NPKCa, chemical nitrogen, phosphorus, and potassium plus lime; NPKM, NPK plus swine manure; and M, swine manure. The results of 29Si NMR spectra support the presence of nanominerals in organic (i.e.,

Selective extraction also showed that the Al fractions were significantly (*P <* 0.05) altered by the long-term fertilization treatment (**Figure 10**). Significantly higher amorphous Al concen‐ trations among the fertilizers were ranked as M ≈ NPKM > NPK ≈ NPKCa > N > NCa > Control (**Figure 10**). Significantly higher strongly organically bound Al concentrations ranked as NPKM > Control > NCa > NPK > NPKCa > M > N; significantly higher weakly organically

NPKM and M) rather than chemical (i.e., N, NCa, NPK, and NPKCa) fertilization treatments.

observed by HRTEM images of soil DOM. [4]

16 Organic Fertilizers - From Basic Concepts to Applied Outcomes

**Figure 15.** Aluminum fractions in the different fertilization treatments from the site of the long-term fertilization ex‐ periment, obtained by selective dissolution techniques. [28] Significant differences between fertilization treatments were determined using one-way ANOVA followed by Duncan's multiple range test at *P* < 0.05, in which conditions of normality and homogeneity of variance were met. The data are shown as mean ± SD (*n* = 3). Control, no fertilization; N, chemical nitrogen; NCa, chemical nitrogen plus lime; NPK, chemical nitrogen, phosphorus, and potassium; NPKCa, chemical nitrogen, phosphorus, and potassium plus lime; NPKM, NPK plus swine manure; M, swine manure.

Meanwhile, Fe K-edge X-ray absorption fine structure spectroscopy (XAFS) is used for both identification and quantification of different mineral phases present in soil colloids.[5,29] Linear combination fitting (LCF) of soil colloids (**Figure 16** and **Table 6**) showed that goethite (56.8–67.0%) and hematite (14.9–25.0%) were prominent under all three fertilizations. The remaining Fe phases were composed of the less crystalline ferrihydrite species. The percentage of ferrihydrite was the highest under NPKM (18.0 ± 0.02%), followed by Control (16.0 ± 0.03%) and NPK (6.30 ± 0.02%). In view of the better C binding and potential preservation capability of ferrihydrite when compared to goethite and hematite,[5,30-32]. Fe minerals under organic fertilization should have a greater C loading than chemical fertilization.

**Figure 16.** Fe K-edge XANES spectra of reference materials and soil colloids from three contrasting long-term (1990– 2014) fertilization treatments. [29] The scattered circles represent the linear combination fitting (LCF) results of the sample spectra. Control, no fertilization; NPK, chemical nitrogen, phosphorus, and potassium fertilization; NPKM, chemical NPK plus swine manure fertilization.


Note: Control, no fertilization; NPK, chemical nitrogen, phosphorus, and potassium fertilization; NPKM, chemical NPK plus swine manure fertilization; ND, not detected. Determination of parameters of fit (i.e., *R*-factor and Chisquare) indicated that the LCF results are convincing.

**Table 6.** Linear combination fit (LCF) results of Fe K-edge XANES spectra of the soil colloids from three separate longterm (1990–2014) fertilization treatments [29]

Nanoscale secondary ion mass spectrometry (NanoSIMS) has the potential to examine the spatial integrity of soil microenvironments and has been designed for high lateral resolution (down to 50 nm) imaging, while still maintaining high mass resolution and high sensitivity (mg kg−1 range).[5,33]. NanoSIMS images, combined with the region of interests (ROIs) analysis, were used to explore the C-binding capability of Al and Fe minerals. Based on the pixel value of secondary <sup>12</sup>C<sup>−</sup> ion mass in all spots from each sample, the selected ROIs were identified. The selected ROIs were further divided into <sup>12</sup>C<sup>−</sup> -rich and <sup>12</sup>C<sup>−</sup> less-rich ROIs. The area percentage of the <sup>12</sup>C<sup>−</sup> -rich or <sup>12</sup>C<sup>−</sup> less-rich ROIs accounted for 7.47 or 40.18%, 10.80 or 27.64%, and 8.23 or 37.99% under Control, NPK, and NPKM, respectively. Interestingly, the box plots (**Figure 17**) of <sup>12</sup>C<sup>−</sup> / <sup>27</sup>Al<sup>16</sup>O<sup>−</sup> (a, b) and <sup>12</sup>C<sup>−</sup> / <sup>56</sup>Fe<sup>16</sup>O<sup>−</sup> (c, d) ratios showed that both the median and the mean values were higher under NPKM than under NPK. These results suggest that Al and Fe minerals under NPKM can bind more organic C than those of NPK.

**Figure 16.** Fe K-edge XANES spectra of reference materials and soil colloids from three contrasting long-term (1990– 2014) fertilization treatments. [29] The scattered circles represent the linear combination fitting (LCF) results of the sample spectra. Control, no fertilization; NPK, chemical nitrogen, phosphorus, and potassium fertilization; NPKM,

**Treatment LCF results (%) LCF parameters**

**Control** 66.0 ± 0.025 14.9 ± 0.000 16.0 ± 0.025 ND 3.10 ± 0.012 ND 0.000052 0.00437 **NPK** 67.0 ± 0.025 25.0 ± 0.000 6.30 ± 0.020 ND ND 1.70 ± 0.008 0.000051 0.00426 **NPKM** 56.8 ± 0.025 20.4 ± 0.000 18.0 ± 0.017 4.8 ± 0.018 ND ND 0.000051 0.00436 Note: Control, no fertilization; NPK, chemical nitrogen, phosphorus, and potassium fertilization; NPKM, chemical NPK plus swine manure fertilization; ND, not detected. Determination of parameters of fit (i.e., *R*-factor and Chi-

**Table 6.** Linear combination fit (LCF) results of Fe K-edge XANES spectra of the soil colloids from three separate long-

**Goethite Hematite Ferrihydrite Ferric sulfates Ferrous citrates Ferrous sulfates** *R***-factor Chi-square**

chemical NPK plus swine manure fertilization.

18 Organic Fertilizers - From Basic Concepts to Applied Outcomes

square) indicated that the LCF results are convincing.

term (1990–2014) fertilization treatments [29]

**Figure 17.** Box plots of <sup>12</sup>C<sup>−</sup> / <sup>27</sup>Al<sup>16</sup>O<sup>−</sup> (a, b) and <sup>12</sup>C<sup>−</sup> / <sup>56</sup>Fe<sup>16</sup>O<sup>−</sup> (c, d) ratios reflecting the <sup>12</sup>C<sup>−</sup> rich ROIs (a, c) and <sup>12</sup>C<sup>−</sup> less rich ROIs (b, d) of the soil colloids from three contrasting long-term (1990–2014) fertilization treatments using Nano‐ SIMS (for all spots). [29] Control, no fertilization; NPK, chemical nitrogen, phosphorus, and potassium fertilization; NPKM, chemical NPK plus swine manure fertilization. The 12C<sup>−</sup> rich ROIs include the areas above 90 pixels, and the 12C <sup>−</sup> less rich ROIs include the areas in the range of 90–40 pixels under Control and NPK, which were above 50 pixels, and in the range of 50–30 pixels under NPKM. The number *n* in figures represents the number of the selected ROIs. The line in the middle of the box is the median value and the square in the box is the mean value. The lines that protrude out of the boxes represent the 25th and 75th population percentiles. Outliers are shown as diamonds.

To address the specific C components preserved by reactive minerals, synchrotron-based C 1s near-edge X-ray fine structure (NEXAFS) spectroscopy was used to identify C composition. Compared to NPK treatment, NPKM and M treatments markedly increased carboxylic groups (288.4–289.1 eV) from 24.2 to 33.2% and increased both the aromatic (283.0–286.1 eV) and phenolic (286.2–287.5 eV) groups by greater than 2.8-fold (**Figure 18** and **Table 7**). In conclu‐

sion, organic fertilization treatments (NPKM and M) enhanced the retention of carboxylic and aromatic C by reactive minerals in soils.

**Figure 18.** Organic C composition in the soil colloids from the various long-term fertilization treatments. [6] (a) Con‐ trol, no fertilization; (b) NPK, chemical fertilization; (c) NPKM, chemical plus swine manure fertilization; (d) M, swine manure fertilization.


Control, no fertilization; NPK, chemical fertilization; NPKM, chemical plus swine manure fertilization; M, swine manure fertilization.

**Table 7.** Deconvolution results for using C 1s NEXAFS on soil colloids from the various long-term fertilization treatments [6]
