Effects of One-Time Long-Term Application of Organic–Inorganic Compound Fertilizer on Wheat Photosynthetic Characteristics, Soil Properties and Grain Yield

Abstract

Wheat production demands simplified fertilization strategies to achieve sustainable high yields. This study evaluated the effects of the one-time high-rate application of an organic–inorganic compound fertilizer on soil properties, photosynthetic characteristics, and grain yields. A multi-year field experiment was conducted with a single basal application of an organic–inorganic compound fertilizer at 3600 kg·ha⁻¹, using conventional split chemical fertilization as the control. Compared with the control, this treatment significantly increased soil organic matter and available nutrient content, enhanced the bacteria/fungi ratio, elevated soil enzyme activity, and promoted the conversion of humus into more stable forms. These improvements sustained a higher flag leaf photosynthetic capacity during the grain-filling stage and delayed leaf senescence. The multi-year average grain yield was 5.28% higher than that of conventional split fertilization. The one-time high-rate application of an organic–inorganic compound fertilizer can improve soil biological properties, maintain late-season photosynthetic function, and increase yields, serving as an effective technical measure for simplified, sustainable, and high-yield wheat cultivation in the region.

1. Introduction

Major wheat-producing regions in China have long relied on the split application of chemical fertilizers. This practice not only incurs high labor costs but also leads to soil compaction, reduced microbial diversity, loss of humus, and other issues due to over-application, threatening the sustainable productivity of farmland and food security [1,2]. Although organic fertilizer can improve soil fertility, its low nutrient content necessitates large application volumes to meet crop demands, and the costs of composting and transportation are high, making it economically difficult to promote. Processing organic fertilizer into a pelleted organic–inorganic compound fertilizer can increase nutrient concentration and reduce the costs associated with storage, transport, and application [3]. This type of fertilizer combines the soil-conditioning and fertility-building functions of organic fertilizer with the rapid nutrient supply characteristics of chemical fertilizers. Its organic component can slow down nutrient release, better matching the crop’s demand throughout its growth period; one-time application aligns with the trend towards agricultural simplification and has significant application potential [4].

Currently, research on organic–inorganic compound fertilizers is often based on short-term trials or limited to single-effect evaluations, such as only examining single-season yield responses or changes in a few soil indicators [5], lacking the comprehensive observation of photosynthetic characteristics, soil properties, and yield formation processes under long-term conditions. Regarding photosynthesis, studies have indicated that humic acid in organic fertilizers can increase wheat leaf chlorophyll content and Rubisco enzyme activity [6,7,8], but these studies mostly focus on the early growth stages, and the effect on maintaining photosynthetic function during the grain-filling stage remains unclear. Concerning soil, organic–inorganic compound fertilizers can increase organic carbon input [9], but how soil organic matter transformation affects microbial communities and the nutrient supply capacity after long-term application needs further clarification. Regarding fertilization methods, the one-time application of chemical fertilizers can easily lead to nutrient deficiency in wheat during the late growth stage [10]. Whether organic–inorganic compound fertilizers can overcome this defect and maintain the soil nutrient supply capacity and photosynthetic function over the long term, thereby achieving stable and increased yields, requires verification through long-term experiments. From the perspective of crop–soil interaction, the role of organic–inorganic compound fertilizers is not merely to substitute part of the chemical fertilizer but rather to promote sustained nutrient release through organic carbon input, humification, and microbial activation, thereby influencing photosynthetic maintenance and yield formation during the late growth stage of wheat. Previous studies have shown that partially replacing chemical fertilizer with organic fertilizer can increase soil organic matter, nutrient content, and microbial activity in wheat fields and improve yield components [11,12,13]. Whether single or slow-release fertilization can achieve stable yields depends on the degree of matching between nutrient release and the wheat demand in the mid-to-late growth period [14,15,16]. Therefore, the key scientific question concerning the long-term single application of an organic–inorganic compound fertilizer is whether it can continuously improve the soil nutrient-supplying capacity, maintain the photosynthetic function of flag leaves during the grain-filling stage, and promote yield formation while reducing the number of fertilizer applications.

Therefore, relying on a 14-year field positioning experiment, this study established four treatments: no fertilizer, conventional split chemical fertilizer, and one-time application of low and high rates of an organic–inorganic compound fertilizer. The effects of each treatment on wheat flag leaf photosynthetic characteristics, soil physicochemical and biological properties, and yields were systematically analyzed to clarify the comprehensive effects of the long-term, one-time application of an organic–inorganic compound fertilizer, providing a technical basis for simplified, high-yield wheat cultivation in the region. On this basis, this study proposes the hypothesis that the long-term single application of an organic–inorganic compound fertilizer can improve the soil physicochemical and biological properties, enhance the nutrient-supplying capacity in the late growth stage, delay the photosynthetic decline of flag leaves, and ultimately achieve stable and increased yields.

2. Materials and Methods

2.1. Study Site and Experimental Design

This experiment began in October 2010. The experimental field was located in Shaomaying Village, Dezhou City, Shandong Province (115°45′ to 117°24′ E, 36°24′ to 38°0′ N). This area is situated in the Yellow River alluvial plain, where non-calcareous fluvo-aquic soils have developed. It belongs to a typical temperate continental monsoon climate zone. Local climate data show an average annual temperature of 12.9 °C and average annual precipitation of approximately 547.5 mm.

A typical winter wheat–summer maize rotation system is adopted. The winter wheat variety is the high-yield, high-quality cultivar Jimai 22, and the summer maize variety is the widely planted Zhengdan 958. Before the experiment, the basic soil nutrient status was systematically measured: the soil pH was 8.3 (weakly alkaline); the organic matter content was 20.3 g·kg⁻¹; the total nitrogen, total phosphorus, and total potassium content was 1.2 g·kg⁻¹, 0.83 g·kg⁻¹, and 20.8 g·kg⁻¹, respectively; the available phosphorus content was 37.4 mg·kg⁻¹; and the available potassium content was 282 mg·kg⁻¹. This provided detailed soil background data for subsequent research.

2.2. Experimental Design

2.2.1. Experimental Treatments

A randomized block design was used with four treatments, each replicated four times, totaling 16 plots. The area of a single plot was 50 m² (10 m × 5 m). The specific treatments were as follows: CK—control with no fertilizer application; CF—conventional nitrogen, phosphorus, and potassium fertilizer treatment; OF1—application of 1800 kg·ha⁻¹ pig manure organic–inorganic compound fertilizer; OF2—application of 3600 kg·ha⁻¹ pig manure organic–inorganic compound fertilizer.

All experimental plots remained fixed with consistent boundaries and corresponded one-to-one between wheat and maize seasons, ensuring the continuity and uniformity of the long-term fertilization treatments. The fertilization regimes for maize were matched to those for wheat, as described below.

(1) Conventional NPK control treatment (CF): The total nutrient input in the maize season was 250 kg ha⁻¹ pure N, 45 kg ha⁻¹ P₂O₅, and 45 kg ha⁻¹ K₂O. All phosphate and potash fertilizers were applied once as a basal fertilizer. Urea served as the nitrogen source, with 1.09 kg applied as a basal fertilizer per plot and an additional 1.63 kg topdressed at the jointing stage, thus splitting nitrogen into two applications. In the wheat season, the CF treatment received total nutrients of 300 kg ha⁻¹ pure N, 120 kg ha⁻¹ P₂O₅, and 100 kg ha⁻¹ K₂O. For each plot, 1.3 kg urea was applied as a basal fertilizer, followed by two topdressings of 0.98 kg urea at the regreening and jointing stages, respectively; all phosphate and potash fertilizers were incorporated into the basal fertilizer.

(2) Pig manure-based organic–inorganic compound fertilizer treatments (OF1 and OF2): The fertilization pattern for maize was identical to that for wheat. OF1 supplied total nutrients of 162 kg ha⁻¹ pure N, 54 kg ha⁻¹ P₂O₅, and 54 kg ha⁻¹ K₂O, whereas OF2 provided 324 kg ha⁻¹ pure N, 108 kg ha⁻¹ P₂O₅, and 108 kg ha⁻¹ K₂O. The full amount of the compound fertilizer was incorporated as a one-time basal application before sowing, with no supplementary topdressing throughout the growing season. All agronomic measures were identical across plots, except for the fertilization treatments. Unified field management protocols were strictly implemented over the 14-year experimental period, guaranteeing the stability and repeatability of the experimental results. The fertilizers used included urea (46% N), triple superphosphate (46% P₂O₅), and potassium sulfate (50% K₂O). The pig manure organic–inorganic compound fertilizer (total N + P₂O₅ + K₂O ≥ 15%, N-P₂O₅-K₂O ratio: 9-3-3, organic matter ≥ 20%, beneficial bacteria 3 × 10⁸ to 5 × 10⁸ per gram) was produced and supplied by Jiangyin Lianye Biotechnology Co., Ltd. Specific fertilizer application rates are detailed in Table 1.

Table 1. Fertilizer application rates for each treatment (kg·ha⁻¹).

Treatment	Urea	Calcium Superphosphate	Potassium Sulfate	Pig Manure Organic–Inorganic Compound Fertilizer
CK	0.00	0.00	0.00	0.00
CF	652.17	260.87	200.00	0.00
OF1	0.00	0.00	0.00	1800
OF2	0.00	0.00	0.00	3600

2.2.2. Experimental Period

The positioning experiment started in October 2010 and continued until June 2024, covering 14 wheat growth cycles. The data in this study were derived from the experimental results for 2022–2024 (seasons 12–14), representing the stable effects after long-term positioning application.

The tested wheat variety was “Jimai 22”. Sowing occurred in mid-October each year, and harvest was in mid-June of the following year. The sowing rate was 150 kg·ha⁻¹, row spacing was 20 cm, and the planting density was approximately 2.25 million plants·ha⁻¹. Except for the different fertilization measures, all other field management practices (e.g., irrigation, pest and disease control) were consistent across treatments.

2.3. Measurement Indicators and Methods

After wheat harvest, soil samples from the 0–20 cm plough layer were collected from each plot using the “five-point plum blossom method” and mixed thoroughly, and plant residues and stones were removed. The samples were divided into two parts: one part of fresh soil was passed through a 2 mm sieve and stored at 4 °C for the determination of the soil microbial quantity and enzyme activity; the other part was air-dried, ground, and sieved for the analysis of soil nutrients [17] and humus components. The soil samples were collected using the same method each year, and all soil and physiological indicators measured were obtained from the same growth stage.

2.3.1. Determination of Wheat Leaf Photosynthetic Characteristics

At the flowering stage (late April to early May) and the mid-grain-filling stage (20 days after anthesis, around mid-to-late May), on clear, cloudless mornings from 9:30 to 11:30 a.m., an LI-6400 portable photosynthesis system (LI-CORBiosciences, Lincoln, NE, USA) was used to measure the net photosynthetic rate (Pn), stomatal conductance (Gs), intercellular CO₂ concentration (Ci), and transpiration rate (Tr) of wheat flag leaves. Photosynthetic parameters were also measured using the LI-6400 portable photosynthesis system (LI-COR Biosciences, Lincoln, NE, USA). The target photosynthetic photon flux density (PPFD) inside the leaf chamber was set to 1000 μmol·m⁻²·s⁻¹ and precisely regulated by the built-in LED light source of the instrument, with actual measured values ranging from 999 to 1001 μmol·m⁻²·s⁻¹. The CO₂ concentration in the leaf chamber was set to the automatic control mode, and the reference CO₂ concentration was maintained at 430–453 μmol·mol⁻¹ to match the ambient atmospheric CO₂ concentration in the field. A constant leaf–air temperature difference control mode was adopted for leaf temperature regulation. The leaf chamber temperature control system stabilized the temperature difference between the leaf chamber and the field ambient environment, and a leaf temperature sensor was equipped to monitor the real-time leaf temperature, which remained between 24.9 and 29.4 °C during measurement. Although all measurements were carried out under open-field outdoor conditions, the closed-loop regulation of the light intensity, CO₂ concentration, and leaf temperature by the instrument effectively minimized interference from external environmental fluctuations, thus guaranteeing the reliability of the measured data and the repeatability of this study.

Ten uniformly growing plants were selected from each plot, and one fully expanded flag leaf was measured per plant. Simultaneously, a SPAD-502 chlorophyll meter was used to measure the relative chlorophyll content (SPAD value) on the same leaves, averaging measurements taken from the upper, middle, and lower parts of each leaf.

The flag leaf area (LA) was measured synchronously at the flowering and mid-filling stages. The leaf length (from ligule to tip) and maximum width were measured with a ruler, and LA was calculated using the following formula: leaf area = length × width × 0.83.

2.3.2. Determination of Soil Enzyme Activity

Fresh-soil chemical colorimetry methods [18] were used to determine the activity of the following enzymes, with three technical replicates and a soil-free blank control for each: urease (URE)—phenol–sodium hypochlorite colorimetric method, with activity expressed as mg NH₄⁺-N released per gram of fresh soil per hour at 37 °C (mg NH₄⁺-N·g⁻¹·h⁻¹); β-glucosidase (β-G)—nitrophenyl-β-D-glucoside colorimetric method, with activity expressed as mg p-nitrophenol released per gram of fresh soil per hour at 37 °C (mg p-nitrophenol·g⁻¹·h⁻¹); cellulase (CL)—3,5-dinitrosalicylic acid colorimetric method, with activity expressed as mg glucose generated per gram of fresh soil over 24 h at 37 °C (mg Glu·g⁻¹·24 h⁻¹); polyphenol oxidase (PPO)—pyrogallol colorimetric method, with activity expressed as mg purpurogallin generated per gram of fresh soil per hour at 30 °C (mg purpurogallin·g⁻¹·h⁻¹); peroxidase (POD)—guaiacol colorimetric method, with activity expressed as mg tetraguaiacol generated per gram of fresh soil per hour at 37 °C (mg tetraguaiacol·g⁻¹·h⁻¹).

2.3.3. Determination of Soil Microbial Quantities

The dilution plate coating method was used to enumerate culturable soil microorganisms [19]. A 10.0 g fresh soil sample was placed in a conical flask containing 90 mL of sterile saline and shaken to prepare a 10⁻¹ soil suspension, followed by serial dilution. Bacteria (B) were cultured on beef extract–peptone agar medium at 28 °C for 3–5 days. Fungi (F) were cultured on Martin’s medium (with rose bengal and streptomycin to inhibit bacterial growth) at 28 °C for 5–7 days. Actinomycetes (A) were cultured on modified Gause’s No. 1 medium (with potassium dichromate to inhibit fungi and bacteria) at 28 °C for 7–10 days. Results were expressed as colony-forming units per gram of dry soil (CFU·g⁻¹).

2.3.4. Determination of Soil Humus Components

Soil humic acid (HA), fulvic acid (FA), and humin (Hm) content was determined using the sodium pyrophosphate–sodium hydroxide extraction–potassium dichromate external heating oxidation methods [20], and the HA/FA ratio was calculated. Air-dried soil samples passing through a 0.25 mm sieve were extracted with a 0.1 mol·L⁻¹ sodium pyrophosphate and 0.1 mol·L⁻¹ sodium hydroxide mixture in a 60 °C water bath with shaking for 2 h. After centrifugation, the total humus extract was obtained. Part of the total extract was acidified to pH 1.0 with 1 mol·L⁻¹ H₂SO₄ and allowed to stand for 24 h to precipitate HA (after centrifugation). The supernatant was FA. The organic carbon content in HA, FA, and the total extract was determined using the potassium dichromate external heating method. The humus component content (g·kg⁻¹ dry soil) was calculated using a conversion factor of 1.724. The HA/FA ratio reflects the degree of humification of soil humus.

2.3.5. Determination of Wheat Yield

From the late dough stage to the harvest of wheat over three consecutive years (2022–2024), yield surveys were conducted in each plot using the five-point sampling method. Five sampling points were set diagonally in each plot. At three representative sampling points per plot, a 1 m² quadrat was established, and all wheat plants within the quadrat were manually harvested, threshed, and cleaned. The threshed grains were naturally air-dried in a ventilated area. The actual moisture content of the grains was measured using a rapid grain moisture meter, and the plot yield was standardized to 13% moisture content.

2.4. Data Statistics and Analysis

Experimental data were first entered, organized, and subjected to basic calculations (mean, standard deviation (SD), standard error (SE), outlier screening) using Microsoft Excel 2021. In this study, indicators including flag leaf photosynthetic characteristics, soil physicochemical properties, and culturable microorganism counts were all measured in the 2023 winter wheat growing season. Therefore, one-way analysis of variance (one-way ANOVA) was adopted to compare differences across different fertilization treatments, and the least significant difference (LSD) method was used for post hoc multiple comparisons. Grain yield data were obtained from three consecutive winter wheat seasons from 2022 to 2024. Accordingly, two-way ANOVA was performed, with the fertilization treatment and year as fixed factors, to analyze the effects of the fertilization treatment, year, and treatment × year interaction on the wheat grain yield. When the treatment effect reached a significant level, the LSD method was further applied for multiple comparisons. The significance level was set at p < 0.05 and the extreme significance level at p < 0.01. Data visualization (bar charts, line graphs, correlation heatmaps, scatter plots, etc.) was performed using the Origin 2021 software. All charts included error bars (standard error (SE)) and were standardized and annotated appropriately.

3. Results

3.1. Effects on Wheat Yield

The box plot (Figure 1) shows that, regarding the yield, the OF2 treatment demonstrated a sustained yield advantage and stability—a key innovative finding. As shown in Figure 1 the OF2 treatment yielded 10.14% and 5.59% higher values than CF in 2022 and 2024, respectively, with an average increase of 5.28%. It showed a continuous optimization trend of increased yields across years. In contrast, although the OF1 treatment showed an advantage in the early stage, its yield decreased in 2024, indicating insufficient stability. The CF treatment maintained stable but limited yield increases.

Figure 1. Wheat grainyieldin 2021–2022 (a), 2022–2023 (b), and 2023–2024 (c) and the 3-year average yield (d) under different fertilization treatments. Different letters indicate significant differences among treatments (p < 0.05).

3.2. Effects on Wheat Leaf Photosynthetic Characteristics

Regarding photosynthetic characteristics, Pn decreased from flowering to grain filling for all treatments (Table 2). OF2 consistently had the highest Pn and the smallest decrease. CF increased Pn by 53.06% compared to CK, while OF1 and OF2 showed successive increases of 7.57% and 8.64% compared to the previous treatment. Gs and Tr showed similar trends, with OF2 maintaining the highest values at both stages. Changes in Ci showed clear treatment differences: Ci significantly decreased for OF1 and OF2 during grain filling, whereas it increased for CF and CK, indicating a stronger CO₂ assimilation capacity for the organic treatments during grain filling.

Table 2. Effects of one-off application of organic–inorganic compound fertilizer on photosynthetic parameters of wheat flag leaf.

Stage	Treatment	Pn/(µmol·m−2·s−1)	Gs/(µmol·m−2·s−1)	Tr/(µmol·m−2·s−1)	Ci/(µmol·m−2·s−1)
Anthesis	CK	15.19 ± 4.06 d	490.62 ± 37.65 c	7.21 ± 0.56 c	313.72 ± 6.20 b
	CF	23.25 ± 0.40 c	558.72 ± 17.85 b	11.65 ± 1.53 a	322.71 ± 4.04 b
	OF1	25.01 ± 0.28 b	530.95 ± 17.61 bc	8.54 ± 0.32 bc	382.20 ± 2.55 a
	OF2	27.17 ± 0.61 a	668.77 ± 13.97 a	9.84 ± 1.27 ab	373.42 ± 9.24 a
Filling	CK	13.23 ± 1.15 b	320.39 ± 9.48 c	5.38 ± 0.89 b	377.52 ± 15.24 a
	CF	18.93 ± 3.26 ab	621.37 ± 7.25 ab	6.97 ± 1.42 a	377.29 ± 16.71 a
	OF1	18.22 ± 1.03 ab	616.39 ± 8.52 b	7.90 ± 2.82 a	371.32 ± 24.13 a
	OF2	19.53 ± 2.65 ab	638.69 ± 8.54 ab	8.06 ± 2.53 a	369.39 ± 4.84 a

Different letters indicate significant differences among treatments (p < 0.05).

Regarding the SPAD values, OF2 had the highest values at both stages (Figure 2a), increasing by 13.46% and 8.76% compared to CF, respectively, indicating a stable photosynthetic material basis. OF1 increased SPAD by 11.44% compared to CF only at anthesis and was comparable to CF at the filling stage. Regarding the leaf area (Figure 2b), OF2 maintained relatively high levels at both the filling and milk stages, increasing by 3.82% and 0.55% compared to CF, respectively, demonstrating both an early light capture ability and sustained photosynthetic capacity in the later stage.

Figure 2. SPAD value (a) and flag leaf area (LA) (b) of wheat under different fertilization treatments at different growth stages. Different letters indicate significant differences among treatments (p < 0.05).

3.3. Effects on Soil Properties

3.3.1. Soil Nutrient Content

The CK treatment had the lowest levels for all indicators, indicating a weak soil fertility base (Table 3). The nutrient reserves and available supply capacity were severely insufficient, with the available phosphorus decreasing by 67.68% and organic matter by 18.13%, resulting in very poor soil support conditions for crop growth. The CF treatment had the highest total nitrogen content, but the increases in available nutrients (except available phosphorus) and organic matter were less than those in OF2, indicating average soil comprehensive fertility stability and sustainability. Compared to CF, the OF1 treatment increased organic matter by 13.75%, provided better available phosphorus and potassium than CK but weaker than CF and OF2, and slightly decreased alkaline hydrolyzable nitrogen by 3.69%. OF1 could serve as a supplementary option for soil fertility improvement. The OF2 treatment had the most significant effect in terms of optimizing soil chemical fertility. Compared to CF, the increases in available phosphorus, available potassium, organic matter, and alkaline hydrolyzable nitrogen reached 33.08%, 10.99%, 23.75%, and 21.59%, respectively. Furthermore, the pH was moderately reduced by 1.47%, potentially activating nutrients with an acidic preference, providing ample, coordinated nutrient supply and a favorable soil environment for crops.

Table 3. Effects of one-off application of organic–inorganic compound fertilizer on soil nutrient content.

Treatment	pH	TN, g·kg−1	AP, mg·kg−1	AK, mg·kg−1	SOM, g·kg−1	AN, mg·kg−1
CK	8.21 ± 0.03 a	0.96 ± 0.09 a	2.56 ± 1.18 b	199.59 ± 35.011 b	13.10 ± 0.21 d	69.07 ± 0.81 c
CF	8.18 ± 0.05 ab	1.16 ± 0.17 a	7.92 ± 2.34 a	225.65 ± 33.97 a	16.00 ± 0.14 c	77.53 ± 6.65 b
OF1	8.16 ± 0.04 ab	1.04 ± 0.07 a	3.87 ± 1.09 b	214.36 ± 21.61 a	18.20 ± 0.15 ab	74.67 ± 4.28 b
OF2	8.06 ± 0.07 b	1.05 ± 0.07 a	10.54 ± 0.98 a	250.46 ± 29.75 a	19.80 ± 0.21 a	94.27 ± 3.52 a

Different letters indicate significant differences among treatments (p < 0.05).

3.3.2. Soil Microbial Community Structure

As shown in Table 4, different fertilization treatments had certain effects on the number of culturable microorganisms in the soil. Compared with CK, the CF treatment increased the numbers of culturable bacteria, fungi, and actinomycetes, indicating that fertilization improved the soil microbial population to some extent. Compared with the CF treatment, the OF2 treatment significantly increased the culturable bacterial count, total microbial count, and bacteria/fungi ratio by 101.9%, 95.9%, and 103.9%, respectively, suggesting that the single application of a high-rate organic–inorganic compound fertilizer is conducive to increasing the number of culturable bacteria and the total microbial population in the soil. The OF1 treatment increased the number of culturable fungi by 13.8% relative to CF, but its enhancement of the total culturable microbial population was relatively limited.

Table 4. Effects of one-off application of organic–inorganic compound fertilizer on soil microbial community structure.

Treatment	B (×105)	F (×103)	A (×104)	TMA (×105)	B/F
CK	13.95 ± 1.99 bc	14.90 ± 2.17 a	5.82 ± 2.42 a	14.68 ± 2.11 b	95.33 ± 22.81 b
CF	17.24 ± 0.91 b	15.47 ± 2.46 a	10.39 ± 1.69 a	18.44 ± 0.99 b	113.44 ± 19.13 b
OF1	15.21 ± 5.03 bc	17.60 ± 5.76 a	7.29 ± 1.85 a	16.11 ± 5.22 b	91.20 ± 31.72 b
OF2	34.81 ± 2.67 a	15.15 ± 1.27 a	11.60 ± 4.83 a	36.12 ± 2.63 a	231.35 ± 32.15 a

Different letters indicate significant differences among treatments (p < 0.05).

3.3.3. Soil Enzyme Activity

Soil enzyme activity varied significantly among treatments (Figure 3). The key finding of this study is that the OF2 treatment fully activated the soil enzyme system. Its cellulase (CEL) and polyphenol oxidase (PPO) activity was significantly higher than that of the CF treatment by 69.62% and 8.14%. In contrast, all enzyme activity in the CK treatment was low. The OF1 treatment exhibited enhanced peroxidase (POD) activity, suggesting that its metabolic pathway leans towards humification processes.

Figure 3. Activity of β-glucosidase (β-GLU) (a), cellulase (CEL) (b), polyphenol oxidase (PPO) (c), urease (UR) (d), and peroxidase (POD) (e) under different fertilization treatments. Different letters indicate significant differences among treatments (p < 0.05).

3.3.4. Soil Humus Composition

Regarding the soil humus composition, the OF2 treatment showed a key advantage (Table 5). Its humic acid content and humic acid/fulvic acid (HA/FA) ratio were significantly higher than those of the CF treatment by 41.8% and 338.5%, respectively. This result indicates that OF2 effectively promotes the transformation of fulvic acid into humic acid, making it the optimal treatment for optimizing the humus composition, enhancing stability, and improving the nutrient retention capacity. In comparison, although the OF1 treatment also increased the humin content and the HA/FA ratio, the extent and pathway of improvement differed from those in OF2.

Table 5. Effects of one-off application of organic–inorganic compound fertilizer on soil humus composition.

Treatment	HA (g·kg−1)	FA (g·kg−1)	Hm (g·kg−1)	HS (g·kg−1)	HA/FA
CK	3.81 ± 1.22 d	3.51 ± 1.27 a	7.36 ± 0.53 c	14.69 ± 0.21 b	1.31 ± 0.97 c
CF	4.81 ± 0.16 c	3.56 ± 0.11 a	9.36 ± 0.68 ab	17.74 ± 0.90 a	1.35 ± 0.02 c
OF1	5.41 ± 0.61 b	1.78 ± 0.15 b	11.22 ± 1.19 a	18.40 ± 0.50 a	3.05 ± 0.30 b
OF2	6.82 ± 0.27 a	1.33 ± 0.49 b	10.34 ± 0.51 a	18.49 ± 0.61 a	5.48 ± 1.50 a

Different letters indicate significant differences among treatments (p < 0.05).

3.4. Correlation and Principal Component Analysis

Correlation analysis showed (Figure 4a) that the wheat yield was significantly positively correlated (p < 0.01) with soil nutrients (total nitrogen, available phosphorus, etc.), microbial indicators (bacteria, actinomycetes, etc.), photosynthetic parameters (Pn, SPAD, etc.), and soil enzyme activity (β-glucosidase, urease, etc.), indicating an interconnected synergistic feedback relationship among these indicators. Principal component analysis showed (Figure 4b) that PC1 contributed 55.5% and PC2 contributed 13.2%. The OF2 treatment performed prominently on positive loading indicators such as yield, soil nutrients, and photosynthetic parameters, distinctly separating it from CK and CF.

Figure 4. Correlation heatmap of wheat yield, photosynthetic characteristics, and soil properties (a) and principal component analysis (PCA) biplot of different fertilization treatments (b). Notes—Y: Yield, TN: Soil Total Nitrogen, AP: Soil Available Phosphorus, AK: Soil Available Potassium, OM: Organic Matter, PH: pH Value, AN: Soil Alkaline Hydrolyzable Nitrogen, B: Soil Bacteria, F: Soil Fungi, A: Soil Actinomycetes, TB: Total Microbial Count, B/F: Bacteria/Fungi Ratio, MBC: Soil Microbial Biomass Carbon, HA: Humic Acid, FA: Fulvic Acid, HM: Humin, HS: Humic Substance, HA/FA: Humic Acid/Fulvic Acid Ratio, β-G: β-Glucosidase, URH: Urease, CL: Cellulase, POX: Polyphenol Oxidase, POD: Peroxidase, Pn1: Photosynthetic Rate at Flowering Stage, Tr1: Transpiration Rate at Flowering Stage, Ci1: Intercellular CO₂ Concentration at Flowering Stage, Gs1: Stomatal Conductance at Flowering Stage, Pn2: Photosynthetic Rate at Filling Stage, Tr2: Transpiration Rate at Filling Stage, Ci2: Intercellular CO₂ Concentration at Filling Stage, Gs2: Stomatal Conductance at Filling Stage, SPAD1: SPAD Value (Chlorophyll Content) at Flowering Stage, SPAD2: SPAD Value (Chlorophyll Content) at Filling Stage, LA1: Leaf Area at Flowering Stage, LA2: Leaf Area at Filling Stage. This figure is a Principal Component Analysis (PCA) biplot. The colored graphical elements are explained as follows: 1. Colored closed ellipses (95% confidence ellipses for each treatment) Each color corresponds to one experimental treatment group. The ellipse represents the 95% confidence distribution range of all samples in the group, which intuitively reflects the intra-group aggregation degree and inter-group differentiation. Black ellipse (CK): Represents the control check group. Samples are concentrated in the left area of the plot. The ellipse is elongated along the PC2 axis, indicating that samples in this group have greater variation on the second principal component. Red ellipse (CF): Represents the conventional fertilization group. It is distributed in the lower-right area near the center, and partially overlaps with the ellipses of CK and OF1, suggesting certain similarities in treatment effects among these groups. Green ellipse (OF1): Represents the first organic fertilizer treatment group. It is located in the upper central area with the largest horizontal span, indicating the highest dispersion of samples in this group on the first principal component. Blue ellipse (OF2): Represents the second organic fertilizer treatment group. It is concentrated in the right area with the most compact shape, meaning the indicators of samples in this group have the highest similarity. Less overlap between ellipses indicates more significant differences in comprehensive indicators between groups. The ellipses of OF2 and CK barely overlap, representing the largest difference among all treatments. 2. Blue arrow lines (variable loading vectors) All blue rays with arrows represent the detected indicators, which explain the driving factors of sample differentiation. Arrow direction: Indicates the positive or negative correlation between the indicator and principal components. Indicators pointing in the same direction are positively correlated, while those in opposite directions are negatively correlated. For instance, POD and PH on the left are positively correlated with each other, and negatively correlated with indicators on the right such as SPAD, SOM and UR. Arrow length: Represents the contribution of the indicator to the principal components. A longer arrow means the indicator has stronger explanatory power for inter-group differences. POD, SPAD, UR and AK are the core indicators driving the grouping differentiation in this plot.

From the variable loadings, indicators such as yield (Y), soil total nitrogen (TN), humus (HS), and humin (HM); crop physiological indicators like the photosynthetic rate at the anthesis and filling stages (Pn1, Pn2) and chlorophyll content (SPAD1, SPAD2); and soil enzymes like β-glucosidase (β-G) and urease (URH) had arrows pointing towards the positive direction of PC1, with high loadings on PC1, playing a key role in distinguishing different fertilization treatments. In contrast, peroxidase (POD) pointed towards the positive direction of PC2, while indicators like pH and fulvic acid (FA) pointed towards the negative direction of PC1, showing different correlations with PC1 and contributions to the principal components.

Overall, the fertilization treatments significantly altered the characteristics of the soil–crop system. Organic substitution treatments (OF1, OF2) differed from CF in their effects on soil humus components, enzyme activity, and crop photosynthetic physiology. For example, organic substitution treatments may enhance crop photosynthetic efficiency and yield formation by improving soil nutrients and enzyme activity. The OF2 treatment was closer to the yield-increasing direction for some indicators, showing a more prominent effect. The PCA visually demonstrates the impacts of different fertilization treatments on the soil–yield system, indicating that the comprehensive effect of organic substitution treatments on soil quality and yields is more significant, providing a visual basis for optimizing fertilization strategies, enhancing soil ecological function, and improving crop yields.

4. Discussion

4.1. Effects of One-Time Application of Organic–Inorganic Compound Fertilizer on Photosynthetic Characteristics of Wheat During Grain Filling

The photosynthetic function of the flag leaf during the grain-filling stage is crucial for wheat grain yield formation [21,22]. This study found that the OF2 treatment maintained higher Pn, Gs, and SPAD values during the grain-filling stage, with the smallest decrease in Pn after anthesis. This benefit is attributed to the sustained nutrient supply characteristic of the organic–inorganic compound fertilizer: its inorganic part can meet the early growth needs of wheat, while the organic part mineralizes slowly, releasing nutrients that coincide with the demand rhythm of wheat during grain filling [23,24], effectively alleviating the premature decline in photosynthetic function caused by late-stage nutrient deficiency associated with conventional chemical fertilizers. Furthermore, the significantly lower intercellular CO₂ concentration (Ci) in the OF2 treatment during grain filling, combined with higher Pn compared to CF, suggests that the maintenance of photosynthetic capacity primarily stems from enhanced mesophyll cell assimilation activity rather than changes in stomatal aperture [25,26].

4.2. Effects of One-Time Application of Organic–Inorganic Compound Fertilizer on Soil Physicochemical and Biological Properties

The soil fertility status is directly influenced by fertilization methods [27]. Under the conditions of this experiment, the OF2 treatment significantly increased the soil organic matter and available nutrient content. This is attributed not only to the direct input of nutrients from the fertilizer itself but also to the improvement of the soil aggregate structure by the organic material, which enhances the water and nutrient retention capacity [28,29,30,31]. Regarding biological properties, the OF2 treatment significantly increased soil bacterial quantities and the bacteria/fungi ratio. A higher bacteria/fungi ratio generally indicates a higher rate of soil nutrient transformation, favoring the conversion of organic nutrients into available forms that crops can directly absorb [32,33]. Simultaneously, the OF2 treatment significantly enhanced the activity of cellulase (related to carbon cycling) and polyphenol oxidase (related to humification) [34,35,36], providing a biochemical basis for sustained soil nutrient supply. Additionally, the substantial increase in humic acid content and the HA/FA ratio under this treatment indicates that the soil humification process tends to form structurally more stable humic acid, which is conducive to the long-term retention and slow release of nutrients [37].

4.3. Analysis of Yield-Increasing Effect of One-Time Application of High-Rate Organic–Inorganic Compound Fertilizer

This study shows that fertilization practices influence wheat yields by altering soil properties, thereby regulating photosynthetic function. Specifically, the long-term application of a high rate of organic–inorganic compound fertilizer (OF2) significantly improved the soil nutrient supply, microbial community structure, and enzyme activity. This favorable soil environment provided a stable nutrient source for wheat throughout its growth period, especially during grain filling, thereby maintaining high flag leaf photosynthetic efficiency [38], delaying plant senescence, and ultimately promoting a grain yield increase (Figure 4).

It is noteworthy that the OF1 treatment did not show a sustained yield-increasing effect in this experiment, indicating a threshold for the application rate of organic–inorganic compound fertilizers. Only when the application rate reached 3600 kg·ha⁻¹ did its soil-improving and yield-increasing effects become consistently apparent, which aligns with previous field trials demonstrating that the yield-promoting effect of organic fertilizers on wheat is often dose-dependent and requires sufficient application rates to achieve stable benefits [39]. Although the CF treatment maintained a certain yield level, its effects in terms of enhancing soil microbial activity and humus quality and delaying photosynthetic function decline were limited. The long-term, single application of chemical fertilizers may lead to the deterioration of the soil physicochemical properties, which is the reason that the one-time application of a high-rate organic–inorganic compound fertilizer showed a comparative advantage in this study [40]. However, it should be noted that, compared with the CF treatment, the OF2 treatment showed differences in total nutrient input, organic carbon input, and the input of organic active substances, so the improvements in relevant traits cannot be fully attributed to the one-time fertilization regime or the organic–inorganic compound fertilizer itself, which is one of the limitations of this study. In addition, the conclusions of this paper only serve to illustrate the effects of the long-term one-time application of an organic–inorganic compound fertilizer on wheat yields, photosynthetic characteristics, and soil properties, and further research on its economic feasibility and promotion value is still required, in combination with fertilizer inputs, labor costs, fertilizer use efficiency and cost–benefit analysis.

5. Conclusions and Prospects

Under the experimental conditions of this study, the one-time application of a high-rate organic–inorganic compound fertilizer (3600 kg·ha⁻¹) improved the soil nutrient status and partial biological properties, as evidenced by the increased soil organic matter and available nutrient content, elevated bacteria/fungi ratio, enhanced activity of related soil enzymes, and promoted transformation of humus into more stable fractions. Meanwhile, the OF2 treatment helped to maintain the high photosynthetic capacity of wheat flag leaves during the grain-filling stage and delayed leaf senescence to a certain extent, resulting in a multi-year average yield that was 5.28% higher than that of the conventional split application of a chemical fertilizer.

Overall, the one-time application of a high-rate organic–inorganic compound fertilizer showed certain effects of soil improvement and yield increase under the experimental conditions; this can provide a reference for the simplified fertilization management of wheat in this region. However, since this study did not evaluate economic benefits, fertilizer use efficiency, environmental risks, or regional adaptability on a larger scale, its practical promotion value still needs further verification. In addition, the impacts of long-term organic–inorganic compound fertilizer application on soil ecological security require further investigation, such as whether organic fertilizer application increases heavy metals or antibiotics in the soil and whether fertilization exacerbates greenhouse gas emissions.