Assessment of Different Humate Ureas on Soil Mineral N Balanced Supply

Abstract

Urea supplements, such as humic acids, could enhance fertilizer nitrogen use effectiveness. Melting is superior to mixing for humate urea application; however, the effects of diverse humate ureas from various coal sources on soil N supply remain unclear. This study compared the properties of two humic acids from different coal sources (HA1, weathered coal; HA2, lignite coal), and their impact on soil mineral N supply and the nitrate–ammonium ratio under flooded and 60% water-filled pore space (WFPS) over a 14-day incubation. Humate ureas stimulated soil mineral N accumulation and balanced the soil nitrate–ammonium ratio at 1:1; however, no significant difference existed between the two humate ureas under 60% WFPS. Humate urea enhanced soil ammonium nitrogen (NH₄⁺-N) retention and delayed nitrate nitrogen (NH₄⁻-N) release, leading to soil mineral N retention, especially in lignite humic acid urea (H2AU) treatments from lignite under flooding. Structural equation modeling (SEM) and linear regression revealed that humic acids elevated soil redox potential (Eh) and electrical conductivity (EC), stimulating soil N mineralization and adjusting the optimal nitrate–ammonium ratio. Humate urea improved soil mineral N supply compared to traditional urea treatments, and humic acids from lignite were more beneficial for crop cultivation from a mineral soil N supply perspective. These findings enhance our understanding of humate urea benefits and aid in optimizing humic acids application for N management.

1. Introduction

Nitrogen (N) is vital for plant growth and crop productivity [1,2]. Exchangeable ammonium (NH₄⁺-N) and nitrate (NO₃⁻-N) are predominant mineral N types assimilated by crops. Research has demonstrated a synergistic effect, indicating that a simultaneous supply of NO₃⁻-N and NH₄⁺-N results in enhanced growth and development indicators, including biomass and dry matter accumulation, compared to single N sources [3,4,5,6,7,8]. The soil nitrate–ammonium ratio hinges on soil N transformation mechanisms, which are significantly influenced by moisture conditions. For instance, Davidson et al. [9] discovered that nitrification and denitrification in arid soil occur concurrently at a soil moisture content of 60~75% WFPS (water-filled pore space); upon exceeding 80–90% WFPS, soil nitrification is restricted [10]. Meanwhile, Faber A et al. showed that a deterioration in water availability led to a 47% decrease in grain yield, a reduction in nitrogen yields, and a 45% decline in nitrogen use efficiency, all under the same fertilization rate (160 ± 40 kg N/ha) [11].

Moisture-specific crops exhibit diverse mineral N-priority patterns. Rice, an ammonium-preferring crop, exhibits optimal growth and photosynthetic characteristics at a nitrate–ammonium ratio of 75:25 during the seedling stage age [7,8]. Xu and Shen [12] also found that the nitrogen nutrient uptake of rice under different water stress conditions was best in the 50:50 and 75:25 treatments. Wheat and tobacco are considered to be ammonium nitrogen-balanced crops, showing significant yield and quality improvement at a nitrate-nitrogen–ammonium ratio of 50:50 [13]. Maize, soybeans, and vegetables, nitrate-preferring crops, experience reduced yield and quality with excessive NH₄⁺-N or excess ammonium-N over nitrate application [14,15]. This suggests that crop preference for N forms may be influenced by the nitrate–ammonium ratio, but the majority of plants respond positively to mixed NO₃⁻-N and NH₄⁺-N.

N uptake by crops primarily originates from soil, mineral fertilizer and natural fertilizers. A 15 N isotope tracer experiment indicated half of crop N absorption originates from soil organic N mineralization, even with fertilizer application [16]. Soil organic N, the primary soil N pool, requires mineralization to inorganic N for plant assimilation [17]. Clearly, soil N supplementation alone is insufficient to meet escalating food demand. Therefore, N fertilizer has been wildly spread around the world, and its usage in agriculture is projected to triple to 249 million tons annually by 2050 [18].

Urea, as the most prevalent N fertilizer, accounts for over 50% of global N-fertilizer consumption [19,20]. When urea is applied in soil, it can be hydrolyzed by urease to form NH₄⁺-N, which can be facilitated by nitrifying bacteria, and can be converted into NO₃⁻-N, both of which are readily absorbed by crops [21,22]. However, excessive urea application can not only decrease crop yield and quality, but also lead to soil acidification, crusting, and a substantial nitrogen loss to the environment through leaching, ammonia volatilization, and surface runoff, posing severe ecological and environmental concerns [23].

Humic acid, a compound formed through the biochemical decomposition of organic matter [24], can stimulate crop growth, regulate soil microbial activity, increase soil organic matter content and humic acid levels [25], adjust soil pH, enhance soil N mineralization and promote N uptake and utilization [26]. As an essential N fertilizer synergist, annually, approximately 350,000 tons of low-grade humic acids are supplemented to urea fertilizers in China [27]. Various methods of humic acid application with urea exist, including blending urea with humic acid. Tian et al. [28] reported that humic acids elevated soil exchangeable NH₄⁺-N and NO₃⁻-N contents. A mineralization incubation study by Zhang et al. [29] found that pre-humic acid inhibited urease activity to reduce ammonia volatilization by 13% and slowed the conversion of NH₄⁺-N to NO₃⁻-N in the soil, thus retaining more soil N. Liang et al. [30] analyzed soil nitrate–ammonium ratio dynamics during wheat growth with urea–lignite-extracted biogenic humic acid, revealing soil nitrate–ammonium ratios of 0.71~0.89 for humic acid and urea co-application treatments at maturity. Mora V et al. [31] found that humic acid promoted the growth of cucumber plant shoots and that high concentrations of humic acid effectively reduced nitrate concentrations in the roots. However, humic acids are powdery, making uniform application challenging.

Recently, melting, as a process combining humic acid and urea hydrolyzed by high-temperature heating, has gained popularity due to its mild reaction conditions, rapid reaction rate, ease of operation, low cost, high yield, and efficiency [32]. For instance, Zhang et al. [21] noted that humic acids derived from weathered coal with high aromatic content could enhance urea-N use efficiency, as their carboxyl groups facilitate the incorporation of -NH₂ into urea, yielding highly stable humate urea compounds that inhibit urea decomposition. Studies have shown that humate urea inhibits nitrification and increases fertilizer N use efficiency by 3.7% to 12% compared to conventional urea [33,34]. Additionally, Yu et al. [35] prepared humate urea from weathered coal using melting, observing that a soil nitrate–ammonium ratio of 1.89 at 5% humic acid addition, which was 9.98% lower than conventional urea, effectively delayed urea hydrolysis. Saha et al. [36] found that lignite–humate urea released only 73% of the added N after 14 days of incubation, compared to traditional urea’s 7-day hydrolysis [37]. This further supports humate urea’s ability to retain more soil N and adjust the soil nitrate–ammonium ratio. Despite studies on the benefits of humate urea by melting, their complex structures from various sources pose challenges in fully understanding their interactions with urea and soil N dynamics.

Given the source, preparation technology, and significant influence of humate urea on soil and fertilizer dynamics, we propose that humate urea can adjust the soil nitrate–ammonium ratio (NAR) to its optimal level (1:1 as standard) via soil N transformation dynamics. We conducted short-term incubations to evaluate the effects of different types of humic acid urea, prepared by the melt process, on changes in soil nitrate–ammonium ratios under two moisture conditions (flooded and 60% water-filled pore space (WFPS)). Additionally, we analyzed the mechanisms underlying these effects. This research aids in selecting the appropriate humate urea type in the field, ultimately contributing to more sustainable agricultural practices.

2. Materials and Methods

2.1. Experimental Soil and Humate Urea Preparation

Soil was procured from the National Field Scientific Observatory of Changshu Farmland Ecosystem, Jiangsu Province, China (31°16′ N, 119°54′ E). The agricultural regimen employed is a rice–wheat rotation system. The primary physical and chemical properties of this tillage layer (0–20 cm) comprised pH 6.17, total nitrogen (TN) 1.41 g kg⁻¹, available phosphorus (AP) 0.41 g kg⁻¹, available potassium (AK) 12.63 g kg⁻¹, and organic carbon (OC) content 11.65 g kg⁻¹. Before experimentation, the soil was naturally air-dried and sieved through a 2 mm sieve to eliminate visible debris.

Humic acids (resistant to acids, soluble in alkali) were extracted from weathered coal and lignite, designated HA1 and HA2, respectively [38,39]. Subsequently, humate urea was synthesized via these steps: initially, 95 g of urea was weighed and heated for final dissolution; secondly, 5 g of humic acid raw material was added and mixed thoroughly using a glass rod; thirdly, it was crushed and ground post cooling to yield humic acid molten urea (HA1U, HA2U). The basic properties of the humic acids are shown in

Table 1. Basic properties of humic acids.

Material	pH	C (%)	N (%)	K (%)	P (%)	Alkyl C (0–45 ppm)	O-alkyl C (45–90 ppm)	Aromatic C (110–160 ppm)	Carbonyl C (160–220 ppm)
HA1	3.8	40.95	0.96	9.8	0.06	15	13.0	50.0	12.0
HA2	3.5	39.13	0.66	10.15	0.02	28.5	17.0	33.0	11.0

.

2.2. Soil Nitrogen Incubation Experiment

A 14-day incubation experiment was instituted at a fixed temperature of 25 °C. Air-dried soil samples were incorporated into cylindrical polystyrene containers (10 cm diameter × 10 cm height), preceded by a 7-day pre-incubation phase with 40% WFPS and 25 °C to restore soil microorganism and enzyme activity. A total of six treatments were carried out: (1) blank soil (CK); (2) urea-only treatment (U); (3) HA1 from weathered coal-only treatment; (4) HA2 from lignite coal-only treatment; (5) weathered coal humic acid urea treatment (HA1U); (6) lignite humic acid urea treatment (HA2U). The control (CK) had no nitrogen input. An instantaneous nitrogen application of 240 kg N ha⁻¹ was coupled with a humic acid raw material routine application of 10 kg ha⁻¹ by the farmers. Upon the uniform mixing of each treatment, the WFPS of each soil sample was adjusted to 60% WFPS under dryland and flooded moisture conditions using deionized water. Each treatment was replicated 3 times. Intense sampling during the formal incubation measured NH₄⁺-N and NO₃⁻-N in the soil on days 0, 1, 3, 5, 7, and 14. Soil total nitrogen, total carbon, pH, soil electrical conductivity (EC) and redox potential (Eh) were assessed on the final day at the end of the incubation.

2.3. Analytical Methods

Soil exchangeable NH₄⁺-N and NO₃⁻ levels were quantified by AA3 continuous flow analyzer (AA3, Skalar, Amsterdam, Holland) [40]. The elemental analysis for TN and OC contents and EC and Eh meter readings for the 1:5 soil–water leachate were conducted using Elementar Vario RL III and Model DDS-11A, respectively; pH was analyzed with a pH meter (S210 Seven Compact™, Mettler, Berlin, Germany) [41]. Solid-state ¹³C NMR spectra of humic acids were performed with a Bruker AVANCE III 400 WB spectrometer (Rheinstetten, Germany) equipped with a contact time of 1 ms and a recycle delay of 5.12 ms. They were normalized to relative areas and grouped into different functional groups according to the expected chemical shift regions for humic acids: alkyl C (0–45 ppm), O-alkyl C (45–90 ppm), aromatic C (90–160 ppm) and carbonyl C (160–200 ppm).

The kinetics of N release was modeled with the popular one-pool exponential equation [42]:

$${\mathrm{N}}_{\mathrm{t}}={\mathrm{N}}_0(1-{\mathrm{e}}^{-\mathrm{K}\mathrm{t}})$$

(1)

where N_t represents the cumulative N mineralized over time (mg kg⁻¹), N₀ indicates the potential mineralized nitrogen potential (mg kg⁻¹); and k denotes the first-order rate coefficient.

The soil N mineralization rate (MIN), the ratio of cumulative soil nitrogen mineralization to total soil nitrogen, was calculated as follows [43]:

$${\mathrm{N}}_{\mathrm{m}\mathrm{i}\mathrm{n}}={\displaystyle \frac{\left({\mathrm{N}\mathrm{H}}_4^{+}-\mathrm{N}+{\mathrm{N}\mathrm{O}}_3^{-}-\mathrm{N}\right)\mathrm{i}-({\mathrm{N}\mathrm{H}}_4^{+}-\mathrm{N}+{\mathrm{N}\mathrm{O}}_3^{-}-\mathrm{N})}{\mathrm{T}\mathrm{N}}}\times 100\mathrm{\%}$$

(2)

where N_min is the soil nitrogen mineralization rate (%), (NH₄⁺-N + NO₃⁻-N)_i is the mineral N post incubation (mg kg⁻¹), NH₄⁺-N + NO₃⁻-N is the mineral nitrogen pre-incubation (mg kg⁻¹), and TN is the TN content of the soil (mg kg⁻¹).

The soil nitrification rate (NIT), the ratio of nitrate nitrogen to mineral nitrogen, was defined as follows:

$${\mathrm{N}}_{\mathrm{n}\mathrm{i}\mathrm{t}}={\displaystyle \frac{{\mathrm{N}\mathrm{O}}_3^{-}-\mathrm{N}}{{\mathrm{N}\mathrm{H}}_4^{+}-\mathrm{N}+{\mathrm{N}\mathrm{O}}_3^{-}-\mathrm{N}}}\times 100\mathrm{\%}$$

(3)

where N_nit is the soil nitrification rate (%), NO₃⁻-N is the nitrate nitrogen content before cultivation (mg kg⁻¹), and NH₄⁺-N + NO₃⁻-N is the soil mineral nitrogen content after cultivation (mg kg⁻¹).

The soil denitrification rate (Denit), the difference between pre- and post-incubation NO₃⁻-N content over the pre-incubation soil nitrate nitrogen content, was calculated as follows:

$${\mathrm{N}}_{\mathrm{d}\mathrm{e}\mathrm{n}\mathrm{i}\mathrm{t}}={\displaystyle \frac{{({\mathrm{N}\mathrm{O}}_3^{-}-\mathrm{N})}_0-{({\mathrm{N}\mathrm{O}}_3^{-}-\mathrm{N})}_{\mathrm{t}}}{{({\mathrm{N}\mathrm{O}}_3^{-}-\mathrm{N})}_0}}\times 100\mathrm{\%}$$

(4)

where N_denit is the soil denitrification rate (%), (NO₃⁻-N)₀ is the soil NO₃⁻-N content pre-cultivation (mg kg⁻¹), and (NO₃⁻-N)_t is the cultivated soil NO₃⁻-N content (mg kg⁻¹).

The soil nitrification inhibition rate [44] can be calculated as follows:

$${\mathrm{N}}_{\mathrm{d}\mathrm{e}\mathrm{n}}={\displaystyle \frac{\mathrm{A}-\mathrm{B}}{\mathrm{A}}}\times 100\mathrm{\%}$$

(5)

where N_den is the rate of inhibition of soil nitrification (%), A is the difference in NO₃⁻-N pre- and post soil incubation in the urea-alone treatment (mg kg⁻¹), and B is the difference in NO₃⁻-N pre- and post soil incubation for the fertilization treatment (mg kg⁻¹).

2.4. Statistical Analysis

Statistical evaluations utilized SPSS 26.0 software. Two-way ANOVA evaluated the influences of soil moisture, fertilizer source, and their interplay on the soil N mineralization rate, nitrification rate, and nitrification inhibition. Linear regression assessed correlations between soil mineral N accumulation and the nitrate–ammonium ratio, with basic soil physicochemical properties (pH, EC, Eh) under varying moisture conditions and adjusted R² as the indicator. IBM SPSS Amos 24.0 Graphics software was employed for structural equation modeling to analyze the effects of various fertilizer treatments on the soil nitrate–ammonium ratio and nitrification inhibition. Mapping was executed using Origin 2021 software.

3. Results

3.1. Comparative Effects of Moisture Conditions on Soil N Supply When Both Humic Acids Are Applied Alone

The two-way ANOVA indicated that soil moisture considerably impacts mineral N accumulation and the nitrate–ammonium ratio with humic acids alone. Specifically, under dryland conditions, there was no significant difference among the CK, HA1, and HA2 treatments (Figure 1). Figure 2 also revealed slight fluctuations in soil NH₄⁺-N and NO₃⁻-N contents among the CK, HA1, and HA2 treatments under dryland conditions. However, soil net mineral N accumulation in HA1 and HA2 treatments was markedly higher (p < 0.05) than that in CK under flooded conditions. Furthermore, humic acids enhanced NO₃⁻-N retention in soil significantly compared to CK and U under flooding (Figure 2). Specifically, the soil NH₄⁺-N content was 4.65 mg kg⁻¹ and 4.45 mg kg⁻¹ in the HA1 and HA2 treatments, respectively, and 43.29 mg kg⁻¹ in the U treatment. Additionally, the U treatment displayed a significantly higher net mineral N content than others under both moisture conditions. Nevertheless, soil nitrate–ammonium ratio dynamics diverged from mineral N accumulation. The soil nitrate–ammonium ratio was significantly higher in humic acid treatments than U under dryland conditions; under flooding conditions, the order of the soil nitrate–ammonium ratio was U > HA2 > HA1 > CK. These findings suggest that HA2 application is advantageous for attaining a nitrate–ammonium ratio of 1:1.

3.2. Humate Urea Application Promote Soil Mineral N Accumulation and Adjust Nitrate–Ammonium Ratio under Dryland Condition

Humate urea notably augmented the net soil mineral N during incubation under dryland conditions. The soil nitrate–ammonium ratio was above one and was indistinctly varied among these treatments (Figure 3). Upon conclusion, the HA1U and HA2U treatments reached 37.36 mg kg⁻¹ and 41.41 mg kg⁻¹ under 60% WFPS; there was no significant distinction between humate urea treatments and conventional urea treatments (Figure 4).

Table 2. Model parameter for cumulative mineral nitrogen in soil incubated for 14 days.

Moisture	Treatment	N0 (mg kg−1)	K (mg N kg−1 Soil Day−1)	R2
Dryland	U	58.32	1.37	0.72
	HA1U	51.34	1.12	0.93
	HA2U	68.53	1.07	0.78
Flooded	U	34.03	1.58	0.65
	HA1U	38.84	1.82	0.72
	HA2U	51.57	1.75	0.83
Moisture		***	***	**
Treatment		**	p > 0.05	***
Moisture*Treatment		p > 0.05	p > 0.05	p > 0.05

Notes: ** and *** indicate significant correlations at 0.05, 0.01 and 0.001 levels, respectively.

shows that HA2U (68.53 mg kg⁻¹) noticeably elevated soil N₀ contents compared with the HA1U (51.34 mg kg⁻¹) and U treatments (58.32 mg kg⁻¹). Humate urea notably (p < 0.05) enhanced the soil mineralization rate. The soil nitrification rate was arranged as follows: HA2U (−147.87%) < U (−109.40%) < HA1U (−72.93%), respectively. Figure 5 also denotes HA1, HA2, HA1U and HA2U diminished soil nitrification suppression efficiency. Linear regression also indicated significant (p < 0.001) positive correlations between soil NH₄⁺-N, NO₃⁻-N, MIN and soil EC and Eh, excluding soil pH. Contrasting results were found between soil EC, Eh and the nitrate–ammonium ratio. These findings suggest that humate urea stimulates soil mineral N accumulation and adjusts the soil nitrate–ammonium ratio to 1:1; however, there is no significant difference between the two humate urea types.

3.3. Humate Urea Extracted from Lignite Promote Soil Mineral N Accumulation by Inhibiting Nitrification under Flooded Condition

Flooded incubation enhanced net soil mineral N with humate urea. The soil nitrate–ammonium ratio was lower than 1, and it varied negligibly among treatments (Figure 3). As illustrated in Figure 4, the soil NH₄⁺-N concentration of HA1U and HA2U treatments reached 50.16 and 70.39 mg kg⁻¹, respectively, under flooding, surpassing the U treatment (43.29 mg kg⁻¹). The soil NO₃⁻-N content differed significantly between the humate urea and traditional urea treatments during incubation, reflecting soil N transformation dynamics. At the end of the incubation, the soil NO₃⁻-N content of the U, HA1U, and HA2U treatments was 4.72 mg kg⁻¹, 6.40 mg kg⁻¹, and 5.61 mg kg⁻¹. Humate urea elevated the soil N mineralization potential as per the soil N mineralization model (Table 2). However, soil nitrification and denitrification rates remained insignificant among the U, HAU1, and HAU2 treatments. Significantly (p < 0.05), HA1U and HA2U enhanced soil nitrification inhibition efficiency compared to the HA1 and HA2 treatments (Figure 5). A significant (p < 0.001) negative correlation was observed between soil pH and soil NH₄⁺-N and mineral N content (Figure 6). Contrary results were obtained for the relationship between soil NH₄⁺-N, mineral N, and soil EC and Eh. In summary, humate urea promotes soil NH₄⁺-N retention and delays NO₃⁻-N release, resulting in soil mineral N retention, particularly in HAU2 treatments from lignite.

4. Discussion

4.1. Humic Acids from Lignite Promote Soil Mineral N and Optimal Nitrate–Ammonium Ratio

Humic acetic acids, crucial N fertilizer synergists [26,30], are assessed for their influence on soil mineral N supply and the nitrate–ammonium ratio balance under flooded and 60% water-filled pore space (WFPS) conditions through an incubation trial. Regarding negligible differences in soil net mineral N and the nitrate–ammonium ratio between the two humic acid treatments, regardless of urea addition (Figure 1 and Figure 3), the reason might be attributed to the low addition of humic acids. The humic acids notably stimulated soil nitrification compared to the urea-alone treatment during the incubation (Figure 5). Structural equation modeling (SEM) elucidated the effects of diverse humic acids on the soil nitrate–ammonium ratio and nitrification inhibition rates under drylands and flooded conditions. The total values of the factors were 95% and 41% in drylands; the total values of the factors were 95% and 71% under flooding, respectively (Figure 7).

Standardized total effects indicated that urea application alone, contrasting with humic acids, reduced soil pH and TOC, hindered nitrification and denitrification processes, and decreased the soil nitrate–ammonium ratio in drylands (Figure 7). But the soil nitrate–ammonium ratio of these treatments was higher in the 1:1 standard (Figure 3). The decreased soil nitrate–ammonium ratio in the U treatment might be attributed to the decreased soil N retention and increased soil N loss. Compared with the U treatment, the effects of different humic acids on the soil nitrate–ammonium ratio varied. HA1 significantly elevated the soil Eh content under the 60% WFPS condition (Figure 7). The concentration of soil Eh content was significantly positively correlated with the soil N denitrification rate. This may be attributed to the high aromatic C (stable C) in HA1. Previous research has shown that bacteria with aromatic hydrocarbon degradation abilities, such as Thauera, facilitate denitrification when fewer alkane hydrocarbons and more aromatic hydrocarbons are present [45]. In addition, HA1 contains a low level of alkyl carbon, which is resistant to degradation and accumulates in the soil. This accumulation can exacerbate soil nitrification, reducing the efficiency of nitrification inhibition and increasing the soil nitrate–nitrogen ratio (Figure 7). However, the positive effects of HA2 on the soil N mineralization rate were superior to those of HA1. HA2 increased the soil Eh and EC, leading to the augmentation of the soil N mineralization rate and denitrification rate, resulting in an decrease in the soil nitrate–ammonium ratio. As Figure 6 illustrates, increased soil EC led to an increase in soil mineral N accumulation. Given that the ideal nitrate–ammonium ratio for crops in arid regions is nitrate–ammonium equilibrium and nitrate-biased nitrate-preferring [13], we believed that HA2 from lignite is advantageous for soil EC increase, resulting in an augmented soil N mineralization rate and adjusting the soil nitrate–ammonium ratio close to the 1:1 standard under the dryland condition.

Contrarily, urea alone application decreased soil pH and soil N nitrification rate, leading to the soil increased nitrate–ammonium ratio under flooded condition. Although Figure 1 and 3 demonstrated the soil nitrate–ammonium ratio was significantly lower than 1:1 under flooded condition, the low soil mineral N cannot be ignored in U treatment. Thus, we need to find the method to elevate the soil mineral N and nitrate–ammonium ratio. As Figure 7 illustrated, HA2 also significantly increased the soil N mineralization rate, N nitrification inhibition efficiency and nitrate–ammonium ratio, which is optimal for the soil nitrate–ammonium ratio under the flooded condition. It was consistent with that under dryland condition. However, soil N denitrification rate in HA2 under flooding differed compared with that in dryland condition. Moreover, increased soil EC also promoted the soil N mineralization rate (Figure 7). The effects of HA1 on soil N transformation dynamics were lower than that in HA2. Given that the ideal nitrate–ammonium ratio for crops in flooded regions is nitrate–ammonium equilibrium and nitrate-bias ammonium-preferring [7,8], we deemed that HA2 from lignite is advantageous for soil EC increase, resulting in the augmented soil N mineralization rate and adjusting the soil nitrate–ammonium ratio close to 1:1 standard under flooded condition.

4.2. Benefits of Humate Urea on Soil Mineral N Accumulation in Diverse Moisture Environments

Soil moisture significantly influences soil mineral N supply and humic acid function. Humate ureas significantly affected soil mineralization parameters, with similar nitrification rates under 60% WFPS and flooded conditions (Table 2 and

Table 3. Analysis of variance of soil N mineralization, nitrification and denitrification rate by different moisture and treatments.

Moisture	Treatment	Soil Mineralization Rate (%)	Soil Nitrification Rate (%)	Soil Denitrification Rate (%)
Dryland	U	2.78 ± 0.22 c	58.94 ± 2.75 a	−109.40 ± 8.26 ab
	HA1U	3.55 ± 0.10 b	58.02 ± 1.86 a	−72.93 ± 10.19 a
	HA2U	4.13 ± 0.15 b	52.99 ± 2.27 a	−147.87 ± 13.86 b
Flooded	U	1.12 ± 0.07 b	9.83 ± 0.87 a	76.86 ± 2.59 a
	HA1U	1.87 ± 0.23 ab	9.79 ± 2.26 a	73.14 ± 6.01 a
	HA2U	2.55 ± 0.94 a	8.10 ± 2.03 a	71.77 ± 2.32 a
Moisture		**	***	***
Treatment		**	p > 0.05	**
Moisture*Treatment		p > 0.05	p > 0.05	**

Notes: **, and *** indicate significant correlations at 0.05, 0.01, and 0.001 levels, respectively. Data are presented as the mean ± standard deviation (n = 3). Different letters indicate significant differences between treatments p < 0.05.

). Linear regression revealed a significant negative correlation between soil pH and soil NH₄⁺-N, NO₃⁻-N and mineral N contents; contrasting relationships were observed for soil EC and Eh under both moisture conditions (Figure 6). This suggests that humate ureas alter soil N dynamics. However, soil net mineral N accumulation varied under both moisture conditions, with HA2U demonstrating superior soil mineral N trends. For instance, HA1U and HA2U enhanced soil mineralization rates by 27.70% and 48.56% under dryland conditions; HA1U and HA2U increased soil mineralization rates by 66.96% and 127.68% under flooded conditions, respectively (Table 3). Notably, humic acids’ minimal N content (Table 1) and their potential contributions to crop N absorption, soil N leaching, and runoff are overlooked due to the constraints of the incubation experiment. Soil mineral N accumulation is influenced by exchangeable NH₄⁺-N and NO₃⁻-N contents, which reflect soil N turnover.

Reeza et al. [46] proposed that humic acids decelerate soil urease activity, thereby reducing NH₄⁺-N release and the mineralization rate. This contradicts our findings. This discrepancy may result from differences in humate urea production methods. Research has examined the effects of humic acids and urea mixture on soil N transformation dynamics [47]; however, our study focused on humic acids bound with urea. Lignite–humate urea exhibits superior soil NH₄⁺-N retention under both moisture conditions (Figure 4). This can be attributed to the high O-alkyl C of HA2, stimulating soil microbial activity and rapid NH₄⁺-N accumulation via soil fixed NH₄⁺-N release [48]. Additionally, humate urea can bind to NH₄⁺-N in the soil biotically or abiotically, with the carboxyl group in the humic acid molecule enhancing NH₄⁺-N retention [49,50]. However, as illustrated in Figure 2 and Figure 4, humic acids alone fail to augment soil NH₄⁺-N content; but humate urea significantly increases NH₄⁺-N content at peak times compared to urea treatment. Urea, an amide N fertilizer, necessitates hydrolysis to generate NH₄⁺-N. Thus, soil NH₄⁺-N increased in humate urea treatments, reflecting the increase in soil urease activity. Urea, an amide N fertilizer, requires hydrolysis to generate NH₄⁺-N. Hence, soil NH₄⁺-N increased in humate urea treatments, indicating increased soil urease activity. This suggests that humate urea may stimulate urea-hydrolyzed NH₄⁺-N rather than soil organic N mineralization. Moreover, melting humate urea may modify the structure of humic acids and enhance their role in soil N ammonization.

Moreover, humate urea may inhibit soil nitrification, slowing the conversion of NH₄⁺-N to NO₃⁻-N, thereby retaining more NH₄⁺-N in the soil. Figure 4 indicated that soil NO₃⁻-N content was lower in humate urea treatments than in the urea-alone application treatment. The results of soil nitrification inhibition efficiency suggested that HA1U and HA2U promoted soil nitrification inhibition under flooded conditions, contrasting with 60% WFPS (Figure 5). This suggests that humate ureas inhibit nitrification, leading to increased soil NH₄⁺-N and mineral N accumulation under flooded conditions compared with 60% WFPS. However, HA1 and HA2 application alone cannot inhibit soil nitrification under both dryland and flooded conditions. This is due to humic acid being a difficult-to-decompose C source, resulting in soil microorganisms needing to absorb more mineral N to meet their energy requirements, promoting soil N assimilation, and the nitrification substrate decreases, slowing soil nitrification [51]. Melting humate urea can modify the structure of humic acids and enhance their role in soil nitrification inhibition. Overall, the results indicate that humate urea is beneficial for soil mineral N accumulation, particularly for lignite–humate urea.

5. Conclusions

This investigation underscores the substantial influence of humic acids on soil N availability during a 14-day incubation, offering insights into their role in augmenting soil mineral N retention and modulating the ideal nitrate–ammonium ratio. Despite its contributions, several limitations must be acknowledged. Our research solely compared soil N dynamics and the nitrate–ammonium ratio between CK, HA1, HA2, HA1U, and HA2U in a brief incubation period. The results showed that the effect of soil moisture on mineral nitrogen accumulation and the nitrate–ammonium ratio varied less with humic acid alone compared to U treatment. Under both moisture conditions, lignite humic acid urea (HA2U) treatment significantly increased soil EC and Eh, decreased soil pH, led to elevated soil mineralization rate, increased soil mineral nitrogen accumulation, and decreased the nitrate–ammonium ratio. Lignite humic acid as a nitrogen fertilizer synergist was superior to weathered coal humic acid for crop cultivation under different moisture conditions. In addition, the study focused on a single soil type and did not explore other indicators of soil quality, such as soil microbial community. Future research should address these gaps by investigating novel methods for humic acid extraction and humate urea production. This will help to better understand the effects of additional C (e.g., humic acids) on soil fertilizer N supplementation an also on soil microbial communities, enzyme activity, and other soil quality indicators. And research needs to continue to assess the shift of nitrogen compounds to plants as this is important in agriculture. Field trials are crucial to validate these findings and promote more judicious fertilization strategies for agricultural productivity.