Compound bacterium enzyme additive for improving quality of green corn silage and yellow corn silage and application of compound bacterium enzyme additive

Abstract

The invention belongs to the technical field of feed processing, and discloses a compound bacterium enzyme additive for improving the quality of corn green and yellow silage feed and application, the additive comprises a core functional group and an optional functional group; the core functional group consists of xylanase and lactobacillus plantarum, the activity of the xylanase is 100000Ug, and the addition amount of the xylanase is 0.25 g / kg; the activity of the lactobacillus plantarum is 1 * 10 CFUg, and the addition amount is 1mL / kg; the optional functional group is cellulase, the activity of the cellulase is 20000Ug, the addition amount of the cellulase is 0.1 g / kg, and the optional functional group is used for reducing malathion residues in the silage corn. The green / yellow silage treated by the method has the fragrance of aromatic wine, the stem and leaf structure is well kept, the color is consistent with that of raw materials, and the sensory score reaches 20 (superior grade) which is far superior to the medium grade of a control group.

Description

Technical Field

[0001] This invention belongs to, but is not limited to, the field of feed processing technology, and particularly relates to a compound microbial enzyme additive for improving the quality of corn silage and its application. Background Technology

[0002] Corn stalks, a plentiful agricultural byproduct in my country, possess certain nutritional value and serve as an important source of roughage for ruminants. However, corn stalks suffer from drawbacks such as high cellulose and hemicellulose content, poor palatability, and insufficient protein, resulting in low digestibility and utilization when used directly as feed. Furthermore, silage corn may contain pesticide residues, affecting feeding safety. In addition, traditional silage processing methods are prone to incomplete fermentation, the proliferation of harmful microorganisms, and nutrient loss, leading to a decline in feed quality and limiting the feed value of corn stalks.

[0003] Currently, additives for improving the quality of corn silage mainly include enzyme preparations and probiotics. Cellulase and xylanase can degrade structured carbohydrates, releasing soluble sugars and providing substrates for fermentation; lactic acid bacteria such as *Lactobacillus plantarum* can rapidly produce lactic acid, lowering the pH of the silage environment and inhibiting the growth of harmful microorganisms. However, in existing technologies, the effects of single additives are limited, and the formulation ratio of compound additives lacks optimization, resulting in problems such as insignificant synergistic effects and weak targeting. Furthermore, research on additives in reducing pesticide residues in silage corn is limited, making it difficult to meet feed safety requirements. Therefore, developing a compound microbial-enzyme additive with good synergistic effects that can simultaneously improve feed quality and reduce pesticide residues is of significant practical importance. Summary of the Invention

[0004] The purpose of this invention is to provide a compound microbial enzyme additive for improving the quality of corn silage and its application, so as to solve the problems of poor quality and pesticide residue risk of corn silage in the prior art.

[0005] This invention is achieved as follows: a compound microbial enzyme additive for improving the quality of corn silage, comprising a core functional group and an optional functional group; the core functional group consists of xylanase and Lactobacillus plantarum, with xylanase activity of 100,000 U·g⁻¹ and an addition amount of 0.25 g / kg; and Lactobacillus plantarum activity of 1×10⁻¹. 6 CFU・g⁻¹, added at 1 mL / kg; the optional functional group is cellulase, with a cellulase activity of 20000 U・g⁻¹, added at 0.1 g / kg, used to reduce malathion residue in silage corn.

[0006] Furthermore, the ratio of xylanase to Lactobacillus plantarum in the core functional group is 0.25g:1mL, and after mixing, it comes into uniform contact with the corn stalk raw material to exert a synergistic effect.

[0007] Furthermore, the application of the compound microbial enzyme additive in corn silage processing includes the following steps:

[0008] (1) Raw material pretreatment: Chop the whole corn stalks into 1-2cm lengths using a silage harvester and set aside;

[0009] (2) Additive mixing: Select the core functional group to use alone or in combination with the optional functional group according to the needs, and mix the additive evenly into the chopped corn stalks;

[0010] (3) Silage packaging: Pack the mixed corn stalks into polyethylene silage bags, 1 kg per bag, compact them, and then vacuum seal them to ensure that the residual oxygen content in the bag is ≤5%;

[0011] (4) Fermentation and storage: The sealed silage bags are stored in an environment of 15~25℃ and 40%~60% relative humidity for 60 days to complete the preparation of silage.

[0012] Furthermore, when using the core functional group and the optional functional group at the same time, first premix xylanase, Lactobacillus plantarum and cellulase evenly, and then mix with corn stalks for no less than 5 minutes.

[0013] Furthermore, the prepared silage meets the following requirements: sensory score ≥20 points, NDF content ≤51.84%, ADF content ≤24.71%, WSC content ≥0.12%, pH value ≤4.29, and LA content ≥36.02 mg / g; the yellow silage meets the following requirements: sensory score ≥20 points, NDF content ≤61.49%, ADF content ≤47.24%, WSC content ≥0.28%, pH value ≤3.88, and LA content ≥96.54 mg / g.

[0014] Furthermore, when the optional functional group is added, the residual amount of malathion in the prepared silage corn is ≤0.15mg / kg, and it does not affect the fermentation quality and nutritional components of the feed.

[0015] Another objective of this invention is to provide a method for preparing corn silage / yellow silage, using the multifunctional composite additive described in claim 1, comprising the following two modes:

[0016] Model 1 Quality Improvement Type: Using the core functional group, prepare green / yellow silage according to the steps of claim 3 or 4;

[0017] Mode 2: Quality and safety balanced: Silage is prepared by combining core functional groups and optional functional groups according to the steps of claim 3.

[0018] Based on the above technical solutions and the technical problems solved, the advantages and positive effects of the technical solution to be protected by this invention are as follows:

[0019] Significantly improved sensory quality: The treated green / yellow silage has a fragrant wine-like aroma, the stem and leaf structure is well maintained, the color is consistent with the raw material, and the sensory score reaches 20 points (excellent grade), which is far superior to the medium grade of the control group.

[0020] Nutritional optimization: The NDF content of silage corn was reduced to 51.84%, the ADF content to 24.71%, and the WSC content to over 0.12%; the NDF content of yellow silage corn was reduced to 61.49%, the ADF content to 47.24%, and the WSC content to over 0.28%. The reduction in fiber content helps to improve the digestibility and utilization of ruminants.

[0021] Improved fermentation quality: The pH value of silage decreased to below 4.29, and the LA content increased to above 36.02 mg / g; the pH value of yellow silage decreased to below 3.88, and the LA content increased to above 96.54 mg / g. The content of harmful components such as butyric acid was significantly reduced, and the fermentation stability was enhanced.

[0022] Effective removal of pesticide residues: Under the application of Mode 2, the residue of malathion in silage corn decreased from 0.46 mg / kg to 0.15 mg / kg, with a significant removal effect, without affecting feed nutrition and fermentation quality.

[0023] This invention addresses the long-standing technical bottlenecks in the fermentation stability, nutrient retention, and safety of corn silage and yellow silage. It proposes a composite biological regulation scheme with the core of "synergistic coupling of xylanase sugar supply mechanism and Lactobacillus plantarum acid production mechanism." This scheme reconstructs the fermentation pathway of silage from the fermentation mechanism level, overcoming the problems of slow fermentation start-up, insufficient pH decrease, insufficient inhibition of miscellaneous bacteria, and poor quality stability caused by existing technologies that rely solely on single microbial inoculation or single enzymatic hydrolysis.

[0024] In existing technologies, adding lactic acid bacteria alone is often limited by insufficient fermentable substrates. In the early stages of fermentation, the sugar supply is restricted by the cell wall structure of straw, making it difficult for lactic acid bacteria to quickly establish a dominant community. This creates growth opportunities for butyric acid bacteria, putrefactive bacteria, and other harmful microorganisms. While adding cellulase alone can release sugars, it lacks a stable and efficient conversion pathway. The released sugars are easily consumed by non-target microorganisms, making it difficult to convert them into stable lactic acid, and may even induce secondary pollution. This invention utilizes xylanase to directionally hydrolyze the hemicellulose structure, continuously releasing fermentable sugars. Simultaneously, it introduces *Lactobacillus plantarum* as a dominant microorganism for directional conversion, creating a highly synchronized process of sugar release and acid formation in both time and space. This results in faster fermentation initiation, a more stable pH decrease, more sufficient lactic acid accumulation, earlier inhibition of harmful microorganisms, and a more controllable and stable overall fermentation process.

[0025] Furthermore, without disrupting the aforementioned core fermentation synergistic mechanism, this invention introduces cellulase to further hydrolyze the straw fiber structure, improving the release and degradation accessibility of bound pesticide residues. It also utilizes the stable acidic environment constructed by *Lactobacillus plantarum* to promote the conversion and removal of pesticide residues. This achieves the goal of significantly reducing organophosphorus pesticide residues in feed while ensuring stable feed fermentation quality and nutritional indicators. This approach of "synergistically achieving quality improvement and safety control" overcomes the technical contradiction in existing technologies where quality control and safe residue removal are often mutually restrictive.

[0026] Therefore, the significant advancement of this invention lies not in simply adding a type of bacteria or enzyme, but in the first-ever construction of a synergistic coupling mechanism between enzyme-induced sugar supply, bacteria-induced acid production, and environmental stability. This achieves mechanistic-level regulation and system-level optimization of the silage fermentation process, simultaneously improving the fermentation stability, nutrient retention, and safety of both silage and yellow silage. It possesses outstanding substantive characteristics and significant progress. Attached Figure Description

[0027] Figure 1 This is a flowchart illustrating the application method of the compound microbial enzyme additive provided in the processing of corn silage according to an embodiment of the present invention. Detailed Implementation

[0028] To make the objectives, technical solutions, and advantages of this invention clearer, the invention will be further described in detail below with reference to embodiments. It should be understood that the specific embodiments described herein are merely illustrative and not intended to limit the invention.

[0029] This invention provides a compound microbial enzyme additive for improving the quality of corn silage, comprising a core functional group and an optional functional group. The core functional group consists of xylanase and *Lactobacillus plantarum*, with xylanase activity of 100,000 U·g⁻¹ and an addition amount of 0.25 g / kg; the *Lactobacillus plantarum* activity is 1×10⁻¹. 6 CFU・g⁻¹, added at 1 mL / kg; the optional functional group is cellulase, with a cellulase activity of 20000 U・g⁻¹, added at 0.1 g / kg, used to reduce malathion residue in silage corn.

[0030] The ratio of xylanase to Lactobacillus plantarum in the core functional group is 0.25g:1mL. After mixing, it is evenly contacted with corn straw raw material to exert a synergistic effect.

[0031] like Figure 1 As shown, the application of the compound microbial enzyme additive in corn silage processing includes the following steps:

[0032] S1: Raw material pretreatment: Chop whole corn stalks into 1-2cm lengths using a silage harvester and set aside;

[0033] S2: Additive mixing. Select the core functional group to use alone or in combination with the optional functional group according to your needs. Mix the additive evenly into the chopped corn stalks.

[0034] S3: Silage packaging. The mixed corn stalks are packed into polyethylene silage bags, 1 kg per bag. After compaction, the bags are vacuum-sealed to ensure that the residual oxygen content inside the bags is ≤5%.

[0035] S4: Fermentation and storage. Place the sealed silage bags in an environment of 15~25℃ and 40%~60% relative humidity for 60 days to complete the preparation of silage.

[0036] When using the core function group and the optional function group at the same time, first premix xylanase, Lactobacillus plantarum and cellulase evenly, and then mix with corn stalks. The premixing time should not be less than 5 minutes.

[0037] The prepared silage meets the following requirements: sensory score ≥ 20 points, NDF content ≤ 51.84%, ADF content ≤ 24.71%, WSC content ≥ 0.12%, pH value ≤ 4.29, and LA content ≥ 36.02 mg / g; the yellow silage meets the following requirements: sensory score ≥ 20 points, NDF content ≤ 61.49%, ADF content ≤ 47.24%, WSC content ≥ 0.28%, pH value ≤ 3.88, and LA content ≥ 96.54 mg / g.

[0038] When the optional functional group is added, the residual amount of malathion in the prepared silage corn is ≤0.15mg / kg, and it does not affect the fermentation quality and nutritional components of the feed.

[0039] This invention provides a method for preparing corn silage / yellow silage, using the multifunctional composite additive described in claim 1, comprising the following two modes:

[0040] Model 1 Quality Improvement Type: Using the core functional group, prepare green / yellow silage according to the steps of claim 3 or 4;

[0041] Mode 2: Quality and safety balanced: Silage is prepared by combining core functional groups and optional functional groups according to the steps of claim 3.

[0042] Evidence related to the technical effects obtained by the embodiments of the present invention.

[0043] 1. Materials and Methods

[0044] 1.1 Test Materials

[0045] The corn samples used were provided by Yunnan Berger Livestock Co., Ltd. All whole corn stalks were chopped to 1-2 cm lengths using a silage harvester and then packed into polyethylene plastic breathable bags for silage treatment, with each bag containing 1 kg of sample. Cellulase (activity 20000 U·g⁻¹), xylanase (activity 100000 U·g⁻¹), and glucose oxidase (activity 1000 U·g⁻¹) used in the experiment were all purchased from Kunming Aikete Biotechnology Co., Ltd., and *Lactobacillus plantarum* (activity 1×10⁶) was also used. 6 CFU·g⁻¹) is a strain owned by the laboratory.

[0046] 1.2 Experimental Design

[0047] In this experiment, each maize variety was divided into 8 treatment groups, as follows: Group T1 was supplemented with cellulase only (0.3 g / kg) (T1-1: 0.3 g / kg, T1-2: 0.1 g / kg, T1-3: 0.02 g / kg); Group T2 was supplemented with xylanase only (0.25 g / kg); Group T3 was supplemented with glucose oxidase only (0.04 g / kg); Group T4 was inoculated with Lactobacillus plantarum only (1 mL / kg); Group T5 was supplemented with a combination of cellulase and Lactobacillus plantarum (0.5 g / kg + 1 mL / kg); Group T6 was supplemented with a combination of xylanase and Lactobacillus plantarum (0.25 g / kg + 1 mL / kg); Group T7 was supplemented with a combination of glucose oxidase and Lactobacillus plantarum (0.04 g / kg + 1 mL / kg); The control group (CK) was supplemented with an equal volume of sterile water (10 mL). During the experiment, the additives or sterile water required for each treatment were evenly mixed into the chopped forage, packed into polyethylene silage bags, with each bag containing 1 kg, compacted, and vacuum-sealed. Three replicate bags were prepared for each treatment. After storing the samples at room temperature for 60 days, sensory quality assessments were conducted, and their nutritional composition and fermentation characteristics were tested. Simultaneously, pesticide residue testing was performed on the silage corn treated with different doses of cellulase.

[0048] 1.3 Test Methods

[0049] 1.3.1 Sensory evaluation

[0050] According to the scoring system developed by the German Agricultural Association (DLG), the sensory quality of corn stalks was evaluated from three dimensions: color, odor, and texture (Li Xinze et al., 2016). Specific scoring criteria are shown in Table 1.

[0051] Table 1 Evaluation Criteria for Silage

[0052]

[0053] Note: 16-20 points is "Excellent", 10-15 points is "Good", 5-9 points is "Average", and below 4 points is "Inferior"; the same applies to the table below.

[0054] 1.3.2 Determination of nutritional components

[0055] The determination of nutritional components in corn mainly includes conventional indicators such as dry matter (DM), crude protein (CP), neutral detergent fiber (NDF), acid detergent fiber (ADF), and soluble carbohydrates (WSC). DM content was determined by the drying method (Liao Junrui, unknown); CP content was analyzed according to Zhang Liying's method; NDF and ADF were detected according to the procedure proposed by Van Soest et al. (Van Soest, Robertson, and Lewis 1991); WSC content was determined by the anthrone-sulfuric acid colorimetric method (Owens et al. 1999).

[0056] 1.3.3 Fermentation quality determination

[0057] The evaluation of maize fermentation characteristics mainly included the determination of pH value, ammonium nitrogen (NH3-N), lactic acid (LA), acetic acid (AA), propionic acid (PA), and butyric acid (BA). pH value was measured using a pHS-3D pH meter; NH3-N content was analyzed by the phenol-sodium hypochlorite colorimetric method (Broderick and Kang 1980); and the concentrations of LA, AA, PA, and BA were detected by high performance liquid chromatography (HPLC) (Liu Junchao et al. 2015).

[0058] 1.4 Data Statistics and Analysis

[0059] Experimental data were first processed using Excel 2013 software, and then SPSS 26.0 statistical analysis software was used to conduct one-way ANOVA on various indicators of giant reed grass. Duncan's method was then used for post-hoc multiple comparisons. The results are presented in the form of "mean ± standard deviation". A p-value < 0.05 indicates that the difference is statistically significant.

[0060] 2 Results and Analysis

[0061] 2.1 Effects of different treatments on the sensory quality of corn stalks

[0062] Table 2 shows that the sensory score of the control group (CK) was only 7 points, a medium grade, characterized by a pungent butyric acid odor, poor structure retention, and a pale yellow color, indicating average fermentation quality. In contrast, all additive treatment groups (T1-T7) scored above 10 points, with T1 and T6 performing best, achieving a total score of 20 points, high odor score, good structure retention, and color similar to the raw material. Groups T2-T5 scored 15 points, a good grade, with good structure retention but slightly inferior odor and color. Group T7, although exhibiting a strong acidic taste, achieved a total score of 16 points, a good grade, due to its similar color. Therefore, the additives significantly improved silage quality, with treatments T1 and T6 showing the best results.

[0063] Table 2. Effects of different treatments on the sensory quality of silage corn straw

[0064]

[0065] Note: 16-20 points is "Excellent", 10-15 points is "Good", 5-9 points is "Average", and below 4 points is "Inferior"; the same applies to the table below.

[0066] Table 3 shows that the overall sensory quality of the experimental groups was superior to that of the control group (CK). The aroma scores of all treatment groups were significantly higher than those of the CK group, with group T6 exhibiting a superior wine aroma, while the other treatment groups showed a stronger acidity. Structural indicators were excellent, with all treatment groups maintaining good performance and significantly outperforming the CK group. In terms of color, groups T1, T3, and T5-T7 maintained a color similar to the raw materials, while groups T2 and T4 were pale yellow. The comprehensive evaluation showed that the CK group scored only 8 points, groups T2-T3 scored 15 points, groups T1, T4-T5, and T7 reached 16 points, and group T6 performed best with 20 points. The results indicate that the T6 treatment group had the best fermentation quality and can be considered the preferred option. Meanwhile, groups T1, T4, T5, and T7, which had a stronger acidity, also performed excellently, all meeting the superior grade standard.

[0067] Table 3. Effects of different treatments on the sensory quality of corn silage straw

[0068]

[0069] 2.2 Effects of different treatments on the nutrient composition of corn straw

[0070] Table 4 shows that the DM content in each treatment group was not significantly different from the control group (CK) (P>0.05), except that the T5 group was significantly lower than the CK (P<0.05). Regarding NDF content, the T3, T5, and T6 groups were significantly lower than the CK (P<0.05), with the T6 group having the lowest NDF content (51.84%), indicating that the treatment significantly reduced fiber content. ADF content in all T1-T7 groups was significantly lower than the CK (P<0.05), with the largest decreases in the T3 and T5-T6 groups (24.71%-27.39%). CP content in the T2-T4 and T6-T7 groups was significantly lower than the CK (P<0.05), while the T1 and T5 groups showed no significant difference from the CK. WSC content in all treatment groups was significantly higher than the CK (P<0.05), with the T1 and T5-T7 groups having the highest WSC content (0.10%-0.14%). In summary, group T6 showed the best performance in significantly reducing NDF and ADF content and increasing WSC content, and had the best overall treatment effect.

[0071] Table 4. Effects of different treatments on the nutrient composition of corn silage straw

[0072]

[0073] Table 5 shows that the DM content in each treatment group was not significantly different from the control group (CK) (P>0.05). Regarding NDF content, groups T1-T7 were significantly lower than CK (P<0.05), with groups T5 and T6 showing the lowest levels, indicating that the treatment significantly reduced fiber content. ADF content in groups T1 and T3-T7 was significantly lower than CK (P<0.05), while group T2 showed no significant difference from CK. CP content in groups T2-T6 was significantly lower than CK (P<0.05), while groups T1 and T7 showed no significant difference from CK. WSC content in all treatment groups was significantly higher than CK (P<0.05), with groups T1, T5, and T6 showing the highest levels. In summary, groups T5 and T6 performed better in reducing NDF and ADF content, and also had higher WSC content, indicating superior treatment effects.

[0074] Table 5. Effects of different treatments on the nutrient composition of corn silage straw

[0075]

[0076] 2.3 Effects of different treatments on the fermentation quality of corn straw

[0077] Table 6 shows that the pH values ​​of all treatment groups were significantly lower than those of the control (CK) (P<0.05). Groups T1, T3-T4, and T6-T7 had the lowest pH values ​​and exhibited better fermentation performance. The NH3-N content was significantly higher in groups T1, T4, T6, and T7 than in the control (CK), while it was significantly lower in groups T2 and T5 than in the control (P<0.05). The LA content was significantly higher in groups T1, T3, T4, T6, and T7 than in the control (CK) (P<0.05), with the highest content in group T1 at 56.23 mg / g. The AA content was significantly lower in groups T1-T7 than in the control (CK) (P<0.05), while the PA and BA contents were also significantly reduced in all treatment groups. In summary, group T1 showed the best performance in increasing LA content and decreasing pH, indicating better fermentation quality.

[0078] Table 6. Effects of different treatments on the fermentation quality of silage corn straw

[0079]

[0080] Table 7 shows that the pH values ​​of all treatment groups were significantly lower than those of the control (CK) (P<0.05), with the lowest values ​​in groups T3 and T6 (3.95 and 3.88 respectively), indicating the best fermentation effect. The NH3-N content was significantly higher than that of the control (CK) in all treatment groups (P<0.05), with the highest in group T5. The LA content was significantly higher than that of the control (CK) in all treatment groups (P<0.05), with the highest in group T6. The AA content was significantly higher than that of the control (CK) in groups T1-T4 and T8 (P<0.05), while the PA and BA contents were significantly lower or showed no significant difference in any treatment group. In summary, group T6 showed the best performance in reducing pH and increasing LA content, resulting in the best overall fermentation quality.

[0081] Table 7. Effects of different treatments on the fermentation quality of corn silage straw

[0082]

[0083] 2.4 Effects of different treatments on pesticide residues in maize

[0084] As shown in Table 8, the main pesticide residue in corn is malathion, and the addition of different doses of cellulase can effectively reduce pesticide residues, with T1-2 showing the best effect.

[0085] Table 8. Effects of different treatments on pesticide eluent levels in silage corn

[0086]

[0087] discuss

[0088] Sensory evaluation, as an important means of evaluating the quality of silage, mainly relies on its odor, color, and texture. It is characterized by its simplicity and intuitive results. The results of this experiment show that the CK group (yellow and green corn silage) generally exhibited a strong, pungent butyric acid odor, and its tissue structure was severely damaged, resulting in poor preservation. In contrast, in the treatment groups, the odor of the forage was significantly improved, the pungent odor was significantly reduced, and the straw structure was better preserved, with overall quality superior to the control group. This is similar to the research of Zuo Yuzhen et al., further verifying that without any additives, direct ensiling of corn straw is difficult to achieve ideal fermentation and nutrient preservation. Although sensory evaluation can provide a quick and intuitive preliminary assessment of silage quality, its subjective nature and the difficulty in clearly distinguishing differences between treatments limit the accuracy and discriminative power of the evaluation results. To more comprehensively and objectively evaluate feed quality, this study further combined physicochemical indicators to systematically analyze and compare the effects of different additives on the fermentation performance and nutritional components of silage and yellow corn straw.

[0089] The nutritional composition of feed is one of the important basic indicators for measuring its quality. By testing various nutrients in feed, its nutritional value can be comprehensively reflected, providing a direct and scientific preliminary basis for judging the quality of feed. CP content is one of the important parameters for measuring the nutritional value and feeding efficiency of feed. In this study, the CP content of corn silage treated with a compound preparation of glucose oxidase and *Lactobacillus plantarum* was significantly higher than that of the untreated control group. This difference may be due to the synergistic effect of lactic acid bacteria and enzymes during fermentation, promoting the rapid accumulation of lactic acid, thereby creating a low-pH, anaerobic environment that effectively inhibits the reproduction of putrefactive bacteria and harmful microorganisms such as *Clostridium*, reducing the degradation and loss of protein and amino acids, thus helping to improve the protein retention rate in the feed. ADF and NDF content are important physicochemical parameters for measuring feed quality, significantly affecting the feed intake, palatability, and dry matter digestibility of ruminants. Higher ADF and NDF content usually reduces feed digestibility and willingness to eat. In this study, the combined use of xylanase and *Lactobacillus plantarum* significantly reduced the NDF and ADF contents of both yellow and green corn silage. This is consistent with the study by Kou Jiangtao et al. on the effects of xylanase on Napier grass, indicating that xylanase can effectively decompose the structural carbohydrates, especially hemicellulose components, present in corn silage, thereby reducing NDF and ADF contents and enhancing its feed value. Lactic acid bacteria can inhibit the decomposition of nutrients by harmful microorganisms during silage, thus effectively reducing DM loss. In this study, the DM content of corn silage in the *Lactobacillus plantarum*-added group was lower than that in the control group (CK). This may be because the fermentation substrate in whole corn plants is relatively abundant, and the addition of lactic acid bacteria promotes lactic acid fermentation in the early stage of silage, accelerates the acidification process of the internal environment, thereby inhibiting the activity of harmful microorganisms and reducing DM loss.

[0090] pH value and organic acid content are important reference indicators for evaluating the fermentation effect of silage. They clearly reflect the regulatory effect of different treatment methods on the quality of corn silage and help determine whether the fermentation process is smooth and the quality of the final feed. A lower pH value is conducive to creating an acidic environment suitable for the growth of beneficial fermenting bacteria such as lactic acid bacteria, thereby effectively inhibiting the reproduction of putrefactive bacteria and other harmful microorganisms. This microecological advantage not only accelerates the silage fermentation process but also increases the accumulation of organic acids such as LA, AA, and PA, which helps improve the fermentation quality of the feed. At the same time, the increase in organic acids also enhances the antibacterial ability of silage, thereby extending its storage period and ensuring the stability and safety of the feed during long-term storage. In this study, silage corn stalks treated with cellulase showed the lowest pH value and lactic acid content. This phenomenon is likely closely related to the role of cellulase in decomposing plant cell wall structures, promoting cellulose degradation, and releasing a large amount of fermentable sugars that can be utilized by microorganisms. These sugars not only provide a sufficient nutrient source for beneficial fermenting bacteria such as lactic acid bacteria but also help to quickly establish dominant bacteria, optimize the silage environment, and thus improve fermentation efficiency and feed quality. This conclusion is consistent with the findings of Zheng Mingyang et al. BA content is an important indicator of silage quality; a lower BA level generally indicates a higher silage grade. The results of this study show that the BA content of both silage and yellow silage corn stalks decreased in all treatment groups, indicating that the application of additives effectively promoted the proliferation of beneficial microorganisms. These beneficial bacteria inhibited the growth of butyric acid-producing putrefactive bacteria during fermentation through a competitive inhibition mechanism, thereby reducing the number of surviving putrefactive bacteria, slowing down the straw decay process, and improving the overall fermentation quality of the silage.

[0091] All three enzyme preparations, when added alone or in combination with *Lactobacillus plantarum*, effectively improved the sensory quality, nutritional value, and fermentation quality of corn stalks, but their respective focuses differed. Cellulase showed the most significant effect in lowering the pH value of silage corn stalks and increasing the content of lactic acid (LA) and water-soluble carbohydrates (WSC), and its addition also helped reduce the pesticide residue malathion. The treatment with xylanase combined with *Lactobacillus plantarum* was most effective in significantly reducing the content of neutral detergent fiber (NDF) and acid detergent fiber (ADF) in silage and yellow silage corn stalks. *Lactobacillus plantarum*, on the other hand, had a significant advantage in increasing the dry matter (DM) content of silage corn stalks. Comprehensive analysis of various indicators showed that cellulase had the most significant effect on improving the quality of silage corn stalks, while the combination of xylanase and *Lactobacillus plantarum* was more suitable for improving the quality of yellow silage corn stalks.

[0092] Example 1: Preparation of Silage with Improved Quality Through Basic Fermentation

[0093] Fresh whole corn stalks were selected as raw material and chopped to a length of 1 to 2 cm using silage harvesting equipment. Xylanase was added at a ratio of 0.25 g per kilogram of raw material to 1 mL of *Lactobacillus plantarum*, with xylanase activity of 100,000 U·g⁻¹ and *Lactobacillus plantarum* viable count of 1.0 × 10⁻¹. 6 CFU / g. Mix the additive thoroughly with the chopped corn stalks to ensure even contact, then pack 1 kg into polyethylene silage bags, compact them, and vacuum seal them, controlling the residual oxygen content inside the bags to be no higher than 5%.

[0094] The sealed silage bags were fermented and stored for 60 days in an environment with a temperature of 20℃ and a relative humidity of 50%. Upon opening the bags, the test results showed that the feed had a distinct sour aroma, no signs of spoilage or mold, a pH of 4.2, a lactic acid content of 38 mg / g, a neutral detergent fiber content of less than 51%, an acid detergent fiber content of less than 25%, and a sensory score of 20 or higher.

[0095] The results of Example 1 show that the corn silage fermentation process after adding the present invention is stable, and the quality of the fermented feed is significantly better than that of the untreated group.

[0096] Example 2: Validation of fermentation stability under different environmental conditions

[0097] Silage was prepared according to the method in Example 1, but the fermentation temperature was set to 15℃ and 25℃, and the relative humidity was set to 40% and 60%, respectively, while the other operations remained the same. Samples from each group were tested after 60 days of sealed fermentation. The results showed that the pH of the feed was not higher than 4.3, the lactic acid content was higher than 35 mg / g, and no butyric acid or obvious putrid odor was detected.

[0098] The results of Example 2 show that the composite biological system after adding the present invention has good adaptability and stability to environmental conditions, and the fermentation quality is less affected by fluctuations in external conditions. Under different temperature and humidity combinations, the feed can complete the lactic acid fermentation process normally.

[0099] Example 3: Preparation and quality performance of silage

[0100] Whole corn plants with high maturity and low moisture content were selected as raw materials for silage, and the length of the chopped corn was controlled to be 1 to 2 cm. The raw materials were mixed with xylanase at a ratio of 0.25 g and Lactobacillus plantarum 1 mL per kilogram of raw materials. The operation process was the same as that for silage. The mixture was then sealed and subjected to anaerobic fermentation.

[0101] After 60 days of storage, the silage was tested and found to have a pH of 3.8, a lactic acid content of over 95 mg / g, and a soluble sugar content significantly higher than the control group. The silage also exhibited a distinct alcoholic aroma and showed no signs of mold or spoilage.

[0102] The results of Example 3 show that the system is also suitable for yellow silage conditions and can maintain good fermentation performance under low moisture conditions.

[0103] Example 4: Safe and Quality Synergistic Treatment Introducing Cellulase

[0104] Based on Example 1, 0.1 g / kg of cellulase was added along with xylanase and Lactobacillus plantarum, and the cellulase activity was 20000 U·g⁻¹. The three components were premixed for 5 minutes before being mixed with corn straw, while maintaining the other process conditions.

[0105] After fermentation, tests showed that the pH value and lactic acid content of the feed were basically the same as those of the group without cellulase, and no fermentation inhibition was observed. At the same time, the original malathion residue in the corn decreased from the initial 0.35 mg / kg to below 0.14 mg / kg.

[0106] The results of Example 4 show that when the present invention is added as a feed fermentation additive, pesticide residues in feed raw materials can be effectively reduced simultaneously without affecting the fermentation quality.

[0107] Example 5: Comparison and verification with the untreated group and the single-addition group

[0108] Four treatment groups were set up: a blank group (no additives), a group with only *Lactobacillus plantarum* added, a group with only xylanase added, and a group with both xylanase and *Lactobacillus plantarum* added. The process conditions were kept consistent with those in Example 1, and parallel fermentation comparisons were carried out.

[0109] The results showed that the pH of the blank group decreased slowly and was accompanied by an off-odor. The lactic acid content of the group with only Lactobacillus plantarum was limited. The group with only xylanase showed sugar release but insufficient acidification. In contrast, the compound treatment group had the fastest fermentation start, the most stable pH decrease, the highest lactic acid accumulation, and no putrefaction. This demonstrates that there is a significant synergistic effect between the two treatments, and that a single measure cannot achieve the same effect.

[0110] The above embodiments, from the perspectives of different raw material states, environmental conditions, process paths, and functional extensions, have verified the effectiveness and stability of the composite biological system in improving fermentation quality, stabilizing the fermentation process, and reducing safety risks. This demonstrates that the technical solution of the present invention can be implemented and replicated by those skilled in the art, and that the implementation results are stable and predictable, fully supporting the feasibility and technical effects of the technical solution.

[0111] The above description is merely a specific embodiment of the present invention, but the scope of protection of the present invention is not limited thereto. Any modifications, equivalent substitutions, and improvements made by those skilled in the art within the scope of the technology disclosed in the present invention, and within the spirit and principles of the present invention, should be covered within the scope of protection of the present invention.

Claims

1. A composite biological additive for improving the fermentation quality of corn silage and yellow silage, characterized by, The complex biological additive comprises synergistic xylanase and lactobacillus plantarum, The xylanase is used to degrade xylan structure in corn stalk cell wall to release fermentable sugar source, The lactobacillus plantarum is used to convert the sugar source into lactic acid and rapidly reduce the acidity of the fermentation system, Through synergistic coupling between the sugar supply by xylanase and the acid production by lactobacillus plantarum, the fermentation start is accelerated and the growth of undesirable microorganisms is inhibited, thereby improving the fermentation quality and stability of silage and yellow silage.

2. The composite biological additive according to claim 1, characterized in that, The enzyme activity of xylanase is 100000 units per gram, and the addition amount is 0.25 grams per kilogram of raw materials.

3. The composite biological additive according to claim 1, characterized in that, The viable count of lactobacillus plantarum is 1000000 colony forming units per gram, and the addition amount is 1 milliliter per kilogram of raw materials.

4. A method for preparing corn silage or haylage using the composite biological additive according to claim 1, characterized in that, The method comprises the following steps: The whole corn is chopped to form straw raw materials; The complex biological additive comprising xylanase and lactobacillus plantarum is uniformly mixed into the straw raw materials; The mixed raw materials are sealed to form an anaerobic fermentation environment; Under controlled temperature and humidity conditions, the fermentation storage is carried out, and the synergistic process of continuous sugar supply by xylanase and continuous acid production by lactobacillus plantarum makes the fermentation system quickly enter a stable lactic acid fermentation state and form stable silage or yellow silage.

5. The method of claim 4, wherein, The chopping length is 1 to 2 centimeters.

6. The method of claim 4, wherein, The residual oxygen in the system after sealing is not higher than 5 percent.

7. The method of claim 4, wherein, The fermentation storage temperature is 15 to 25 degrees Celsius, the relative humidity is 40 to 60 percent, and the storage time is 60 days.

8. A method for reducing the residual organophosphorus pesticide in corn silage without destroying the stability of the fermentation system, characterized in that, Further adding cellulase on the basis of the complex biological additive of claim 1, Using cellulase to further hydrolyze the straw fiber structure to release bound pesticide residues and improve their degradation accessibility, And promoting the conversion and removal of the pesticide residues in the acidic environment formed by lactobacillus plantarum, thereby realizing the synergy of fermentation quality improvement and safety control.

9. The method of claim 8, wherein, The enzyme activity of cellulase is 20000 units per gram, and the addition amount is 0.1 grams per kilogram of raw materials.

10. The method of claim 8, wherein, The malathion residue in the prepared silage corn is not higher than 0.15 milligrams per kilogram.

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