Preparation method and application of nano-selenium compound liquid

Abstract

The invention belongs to the technical field of nano materials and bioengineering, and particularly discloses a preparation method and application of nano-selenium compound liquid, the method comprises the following steps: S1, providing a bacillus licheniformis B1-Se strain, the preservation number of the bacillus licheniformis B1-Se is GDMCCNO: 64806; s2, fermenting the strain in a culture medium containing a carbon source, a nitrogen source and sodium selenite; s3, after fermentation is finished, performing solid-liquid separation on fermentation liquor, and collecting supernate; s4, the supernate obtained in the step S3 serves as a solvent, the pH of the supernate is adjusted to 3.8-4.2, then a dispersing agent, ascorbic acid and sodium selenite are sequentially added for a reaction, and nano-selenium liquid is generated; and S5, adding vitamin E into the nano-selenium liquid obtained in the step S4, and uniformly mixing to obtain the compound liquid. The fermentation supernatant of bacillus licheniformis B1-Se under specific conditions completely replaces pure water to serve as a solvent and a medium for chemical synthesis reaction of nano-selenium, so that the problem of spoilage of the product is solved, and long-acting stability is realized.

Description

Technical Field

[0001] This invention belongs to the field of nanomaterials and bioengineering technology, specifically, it relates to a method for preparing a nano-selenium compound liquid and its application. Background Technology

[0002] Currently, the mainstream production process for nano-selenium liquid products is the surfactant-ascorbic acid (VC) chemical reduction method. This process uses pure water as a solvent and operates under acidic conditions of pH 3.5-4.5, through the redox reaction of sodium selenite and ascorbic acid (reaction formula: 2Na₂SeO₃ + 3C₆H₈O₆ + 4H₂O). + →2Se 0 ↓+ 3C6H6O6 + 4Na + Preparation of red nanoscale elemental selenium (Se) by adding 6H2O 0 This production system relies on 40-60 g / L of surfactant to maintain the dispersion stability of the product. However, this system suffers from two major flaws that severely limit product performance and application scenarios: First, there is a significant risk of spoilage. In high-temperature storage environments, the product's pH value will gradually drift from acidic to neutral (5.0-6.5) over time, creating an environment highly conducive to microbial growth. Especially under high-temperature conditions in summer (>30℃), the product will develop a noticeable putrid odor after 7-15 days of storage, completely rendering it unusable.

[0003] Secondly, it faces the technical challenge of chemical oxidation and blackening. Nanoscale elemental selenium (Se) 0 Exposure to oxygen-containing environments readily leads to a progressive oxidation reaction (Se). 0 →Se 4+ / Se 6+ This caused the product's color to fade from red to black, resulting in a significant decrease in bioactivity. Accelerated testing at 40℃ showed that after only 10 days, the retention rate of the highly bioactive nano-sized elemental selenium was less than 20%, severely impacting the product's efficacy and stability.

[0004] To address the aforementioned shortcomings, various improvement methods have been attempted in related fields, but all have significant limitations. Some solutions inhibit spoilage by adding chemical preservatives such as potassium sorbate, but this can cause intestinal flora imbalance when used in animal feed, violating the core requirements of green farming. Other solutions employ physical protection methods such as silica coating and aseptic nitrogen-filled packaging, which can delay spoilage to some extent, but the former leads to a 300% increase in production costs, and the latter poses a risk of secondary contamination after opening, making both difficult to promote and apply industrially.

[0005] The aforementioned technical solutions all reveal fundamental limitations: various improvement measures only focus on adding exogenous preservatives or physical isolation methods, failing to address the biological defects of the solvent system itself—pure water solvent systems lack both endogenous factors to inhibit microbial proliferation and the ability to establish a micro-ecological balance that resists oxidation. This core technological gap directly restricts the large-scale promotion and application of nano-selenium liquid products in tropical and subtropical regions, becoming a technological bottleneck that the industry urgently needs to overcome.

[0006] In view of this, the present invention is proposed. Summary of the Invention

[0007] The technical problem to be solved by this invention is to overcome the shortcomings of the prior art and provide a method for preparing nano-selenium compound liquid and its application. This method fundamentally improves the micro-ecological environment of traditional nano-selenium liquid by using Bacillus licheniformis fermentation supernatant instead of pure water as the reaction solvent, giving it inherent anti-corrosion and anti-oxidation capabilities, and endowing it with multiple biological functions.

[0008] To achieve the above objectives, the technical solution of the present invention is as follows: In a first aspect, the present invention provides a method for preparing a nano-selenium compound liquid, comprising the following steps: S1. Provide Bacillus licheniformis strain B1-Se, wherein the preservation number of Bacillus licheniformis strain B1-Se is GDMCCNO:64806; S2. Fermentation is carried out using the strain in a culture medium containing a carbon source, a nitrogen source, and sodium selenite; S3. After fermentation, the fermentation broth is separated into solid and liquid components, and the supernatant is collected. S4. Using the supernatant obtained in step S3 as a solvent, adjust its pH to 3.8-4.2, and then add dispersant, ascorbic acid and sodium selenite in sequence to react and generate nano-selenium liquid. S5. Add vitamin E to the nano-selenium liquid obtained in step S4, mix well, and obtain the compound liquid.

[0009] In a preferred embodiment, in step S2: The carbon source is sucrose with a concentration of 25 g / L; The nitrogen source is ammonium sulfate with a concentration of 10 g / L; The concentration of sodium selenite added was 1.1 g / L.

[0010] In a preferred embodiment, citric acid is used to adjust the pH in step S4.

[0011] In a preferred embodiment, the pH is adjusted to 4.0 in step S4.

[0012] In a preferred embodiment, in step S4: The dispersant is chitosan, and the addition amount is 0.0975 kg / t; The amount of ascorbic acid added is 8.25 kg / t; The amount of sodium selenite added resulted in a total selenium content of 0.2% (w / v) in the final product.

[0013] In a preferred embodiment, in step S5: The vitamin E mentioned is modified water-soluble vitamin E with an effective content of 50%, and its concentration added to the final product is 10 kg / t.

[0014] Secondly, the present invention provides a nano-selenium compound liquid prepared by the above method, wherein the nano-selenium compound liquid comprises nano-elemental selenium, fermentation metabolites of Bacillus licheniformis B1-Se, a dispersant, ascorbic acid, and vitamin E.

[0015] In a preferred embodiment, The content of the nano-elemental selenium is 0.2% (w / v); The fermentation metabolites of Bacillus licheniformis B1-Se include antimicrobial lipopeptides and digestive enzymes.

[0016] In a preferred embodiment, the nano-selenium compound liquid further contains live Bacillus licheniformis B1-Se bacteria, with a live bacteria count of not less than 1×10⁻⁶. 6 cfu / mL.

[0017] Thirdly, the present invention also provides the application of the nano-selenium compound liquid as described above in the preparation of animal drinking water additives.

[0018] By adopting the above technical solution, the present invention has the following beneficial effects compared with the prior art: 1. This invention utilizes Bacillus licheniformis fermentation supernatant instead of pure water as the reaction solvent. The Bacillus licheniformis fermentation supernatant itself is rich in natural antimicrobial lipopeptides (such as Surfactin and Fengycin), which create a microenvironment in the system that continuously inhibits the growth of other microorganisms. Simultaneously, the organic acids produced during fermentation, along with citric acid, maintain a stable acidic environment (pH ~4.0). This dual action fundamentally prevents spoilage, achieving stable storage at room temperature for over 90 days without the need for refrigeration or the addition of chemical preservatives, thus solving the problem of product spoilage and achieving long-term stability.

[0019] 2. This invention forms a composite antioxidant system. In addition to vitamin C and the compounded vitamin E, the bacterial metabolites in the fermentation supernatant also contribute additional antioxidant activity. This multi-layered, synergistic antioxidant network can more effectively scavenge free radicals and block oxidation chain reactions, thus maintaining a bright red color for a long time in the accelerated test at 40℃, ensuring the high bioactivity of nano-selenium.

[0020] 3. The nano-selenium compound liquid of this invention can be used to prepare animal drinking water additives, and in addition to selenium supplementation, it also provides: Antibacterial function: The antimicrobial peptides in the supernatant can effectively inhibit intestinal pathogens (such as Escherichia coli and Clostridium perfringens). Digestive aid function: Proteases, amylases, and other enzymes in the supernatant help improve animal digestion; Probiotic function: The product contains live Bacillus licheniformis bacteria (≥10). 6 (CFU / mL) can colonize the gut and continuously exert a probiotic effect; Comprehensive antioxidant protection: alleviates oxidative stress in animals.

[0021] This integrated "nutrition + health care" solution significantly surpasses the value of traditional products and can reduce antibiotic use at the source.

[0022] 4. This invention utilizes fermentation supernatant as a solvent, achieving high-value utilization of waste resources (otherwise, the supernatant would need to be treated as wastewater), thus reducing raw material costs. The entire process is compatible with existing equipment, requiring no huge investment. Furthermore, the product contains no synthetic preservatives, making it safer and more environmentally friendly, aligning with the trends of green farming and antibiotic reduction. Attached Figure Description

[0023] The accompanying drawings, as part of this invention, are provided to further illustrate the invention. The illustrative embodiments and descriptions of the invention are used to explain the invention, but do not constitute an undue limitation thereof. Clearly, the drawings described below are merely some embodiments, and those skilled in the art can obtain other drawings based on these drawings without any creative effort.

[0024] In the attached diagram: Figure 1 This is the growth curve of strain B1-Se under the condition of continuous selenium feeding. Figure 2 This is a schematic diagram showing the changes in bacterial culture under the conditions of continuous selenium feeding of strain B1-Se of the present invention; Figure 3 This is a schematic diagram of the original culture medium and supernatant of strain B1-Se of the present invention via flow-through selenium addition; Figure 4 This is a schematic diagram of the centrifuged bacterial sludge cultured with the B1-Se strain of the present invention via selenium addition; Figure 5 This is a schematic diagram illustrating the stability verification results of the anti-corrosion nano-selenium compound liquid product of this invention; Figure 6 This is a comparative chart of jejunal lesion scores for different groups of broilers according to the present invention; Figure 7 This is a schematic diagram of different groups of jejunal villus slices according to the present invention.

[0025] It should be noted that these accompanying drawings and textual descriptions are not intended to limit the scope of the invention in any way, but rather to illustrate the concept of the invention to those skilled in the art by referring to specific embodiments. Detailed Implementation

[0026] To make the objectives, technical solutions, and advantages of the embodiments of the present invention clearer, the technical solutions in the embodiments will be clearly and completely described below with reference to the accompanying drawings. The following embodiments are used to illustrate the present invention, but are not intended to limit the scope of the present invention.

[0027] [Example 1] This embodiment provides a process for preparing functional fermentation supernatant, which specifically includes the following steps: 1. Strain activation and propagation The preserved Bacillus licheniformis B1-Se strain (GDMCC NO:64806) was inoculated into beef extract peptone liquid medium and cultured at 37°C and 180 rpm for 24 hours to obtain primary seed culture.

[0028] 2. Fermentation culture and biotransformation ( Figure 1 , Figure 2 ) The primary seed culture was transferred to a fermentation medium with a specific composition at a certain inoculation amount (3-5% v / v); Ferment at 35-37℃ with aeration and stirring (150-200 rpm) for 60-72 hours.

[0029] The culture medium used sucrose as the main carbon source (25 g / L) and ammonium sulfate as the main nitrogen source (10 g / L), and added sodium selenite (1.1 g / L) as a selenium source precursor through dynamic feeding.

[0030] 3. Centrifugal separation After fermentation, the fermentation broth is centrifuged at high speed (10,000-12,000 rpm, 15-20 minutes) to achieve solid-liquid separation.

[0031] 4. Product Acquisition ( Figure 3 , Figure 4 ) After centrifugation, red bacterial sludge and amber to light red supernatant were collected separately. This supernatant is the functional fermentation supernatant required in this invention.

[0032] The supernatant contains: a. Antimicrobial lipopeptides produced by the strain (such as Surfactin, Fengycin, etc.). b. Various digestive enzymes (such as protease and amylase); c. Incompletely converted sodium selenite residue (approximately 0.23 g / L); d. Organic acids, polysaccharides, and other active substances produced by bacterial metabolism; e. Viable bacteria count is 10 6 Bacillus licheniformis B1-Se at the cfu / ml level.

[0033] The Bacillus licheniformis strain B1-Se involved in this invention is GDMCC NO:64806. The taxonomic name of this strain is Bacillus licheniformis. It was deposited on June 27, 2024, at the Guangdong Provincial Center for Microbial Culture Collection, located at the Institute of Microbiology, Guangdong Academy of Sciences, 5th Floor, Courtyard 59, No. 100 Xianlie Middle Road, Guangzhou, with accession number GDMCCNo:64806. [Example 2] This embodiment provides the preparation process and stability verification of the anti-corrosion nano-selenium compound liquid. The preparation process is as follows: 1. Provide solvent Take an appropriate amount of the fermentation supernatant prepared in Example 1 as the liquid basis of the reaction system.

[0034] 2. pH adjustment The pH of the supernatant was adjusted to 3.8-4.2, preferably 4.0, using food-grade citric acid solids to create an acidic environment that is suitable for the reduction reaction and inhibits most contaminating bacteria.

[0035] 3. Add reaction raw materials Under stirring conditions, the following were added sequentially to the acidified supernatant: Chitosan 0.0975 kg / t was used as a dispersant for nanoparticles.

[0036] Ascorbic acid (VC) 8.25 kg / t, as the main reducing agent.

[0037] An additional 3.625 kg / t of sodium selenite was added as a supplementary selenium source.

[0038] 4. Reduction reaction The reaction was stirred at 28℃ for 1-2 hours until the solution turned a uniform, transparent, bright red color, indicating that the excess sodium selenite had been completely reduced to nano-elemental selenium (Se). 0 ).

[0039] 5. Compound of active ingredients After the reaction is complete and the system is cooled to room temperature, add 10 kg / t of modified water-soluble vitamin E product (50% effective content).

[0040] 6. Homogenization and Packaging After the final product is stirred evenly, it can be packaged to obtain a bright red anti-corrosion compound liquid product with a nano selenium content of about 0.2%.

[0041] The stability of the anti-corrosion nano-selenium compound liquid was verified as follows: The experimental groups are set up as shown in Table 1 below. After each group is properly set up, the following indicators are observed and detected under the same lighting conditions at the same time each week: a. Directly observe the color and sedimentation level under both static and shaken conditions; b. Use a transparent plastic straw to draw a small amount of liquid in both static and shaken states to observe the grainy texture; c. Odor sensory evaluation (degree of odor production), with a test period of 3 months.

[0042] The stable storage group trial period was 3 months, and the intermittent pouring group trial period was until each bottle was empty according to the pouring procedure.

[0043] Using the simulated intermittent pouring opening procedure: fix the personnel, fix the graduated cylinder (±5 mL), and tighten it within 5 seconds after pouring.

[0044] Result: Reference Figure 5 The first three images from left to right correspond to the images numbered L-Ⅰ-D, L-Ⅰ-B, and L-Ⅰ-C in Table 1, taken up to week 12. The left side of each image shows the surface extract, and the right side shows the deep extract. The results show that, up to week 12, the appearance of groups L-Ⅰ-D and L-Ⅰ-B was not significantly different from that of the fresh product. The liquid in group L-Ⅰ-C (low-temperature refrigeration) became thinner and the color slightly lighter. Furthermore, the fourth image, corresponding to group L-Ⅰ-15, taken on day 90 and after 12 pours, showed no significant difference in liquid appearance from the fresh product. In all four simulations, no liquid stratification or other odors were observed, and the liquid color remained bright red. The conclusion indicates that after 90 days of storage under different conditions, the material maintained good stability regardless of outdoor temperature or light exposure, without any abnormal phenomena such as spoilage, agglomeration, or blackening.

[0045] Table 1. Stability Verification Groups for Liquid Selenium Products serial number selenium concentration Preservation conditions Sample loading conditions Discharge volume & frequency Remark L-Ⅰ-D 0.2% Outdoor temperature, direct sunlight 1L white translucent plastic bottle 0 / L-Ⅰ-B 0.2% Outdoor temperature, blocking sunlight 1L white translucent plastic bottle 0 / L-Ⅰ-C 0.2% 4℃ cold storage 1L white translucent plastic bottle 0 / L-Ⅰ-15 0.2% Outdoor temperature, blocking sunlight 2.5L white translucent plastic bottle 100 ml once a week, pour until finished. Each 100 ml volume was poured out and photographed under the same transparent container and lighting conditions against a black background. The resulting images were then arranged for color change comparison at different times. [Example 3] This embodiment verifies the application effect of nano-selenium compound liquid in broiler waterline feeding. This product, as an animal drinking water additive, possesses multiple functions including selenium supplementation, inhibition of pathogenic bacteria, anti-oxidative stress, and supplementation of digestive enzymes. The verification process is as follows: One hundred and ninety-two healthy AA white-feathered broiler chickens aged 12 days were randomly divided into four treatment groups, with six replicates per group and eight chickens per replicate. The experimental period lasted 42 days, with the first 14 days being a uniform brooding period and the following 35 days being the formal experimental period. The experimental design is as follows: The normal control group (NC group) drank ordinary tap water throughout the process, without adding any selenium source, and did not undergo any challenge.

[0046] Ordinary test group (NS group): 0.1% (v / v) of the nano-selenium compound liquid of this invention was added to drinking water throughout the process.

[0047] The challenge control group (PC group) drank ordinary tap water throughout the entire experiment and underwent a combined challenge with coccidia and Clostridium perfringens on day 21 of the experimental period.

[0048] Treatment group (TR group): 0.1% (v / v) of the nano-selenium compound liquid of the present invention was added to the drinking water throughout the trial, and the group received the same combined challenge as the PC group on day 21 of the trial period.

[0049] The challenge procedure was scheduled for day 21 of the experiment, with chickens in the PC and TR groups orally inoculated with 1 ml of 5×10⁻⁶ dextrose solution. 3 CFU / ml Eimeria tenella oocysts were administered orally on day 22 (1 ml 1×10⁻⁶ oocysts). 9 CFU / ml of Clostridium perfringens type A bacterial suspension. During the experiment, all chickens had free access to feed and water, received routine immunizations, and were managed according to commercial farming standards. Fasting weights were recorded on day 0 (end of brooding) and day 35 (end of the experiment), with replicates, and feed consumption was recorded. On day 25 (day 4 after challenge), four chickens from each group were randomly selected for necropsy, and jejunal mucosal scrapings were performed to observe intestinal lesions and score them (0-4 points, 0 points for no lesions, 4 points for severe lesions). Cecal digesta was also collected for Clostridium perfringens counting.

[0050] After a 21-day trial period, the growth performance, gut health, and disease resistance of the broiler chickens in each group were as follows: 1. Growth performance results As shown in Table 2, at the end of the experiment (day 35), compared with the normal control group (NC), the normal experimental group (NS) with added 0.1% nano selenium compound liquid had significantly higher final body weight and average daily gain (ADG) (P<0.05), and significantly lower feed conversion ratio (FCR) (P<0.05), indicating that this product can effectively promote broiler growth and improve feed utilization under normal feeding conditions.

[0051] The ADG of the challenge control group (PC) was significantly lower than that of the NC group (P<0.05), and the FCR was the highest, indicating that the combined challenge successfully established an intestinal disease model, which had a significant negative impact on growth performance. In contrast, the treatment group (TR), under the same challenge conditions, had significantly better ADG and FCR than the PC group (P<0.05), and no significant difference from the NC group (P>0.05). This indicates that the product of this invention can effectively alleviate the growth inhibition caused by challenge and maintain the normal growth level of broilers.

[0052] Table 2. Effects of nano-selenium compound liquid on the growth performance of broiler chickens Group Initial weight (g) Final weight (g) Average daily weight gain (ADG) (g / animal / day) FCR (Feed to Crop Ratio) NC 420±10 2410 ± 42 a ]] 94.8 ± 1.7 a ]] 1.62 ± 0.03 a ]] NS 422±9 2535 ± 38 b ]] 100.6 ± 1.5 b ]] 1.57 ± 0.02 b ]] PC 418±11 <![CDATA[2275±45 c ]]> <![CDATA[88.4±1.9 c ]]> <![CDATA[1.71±0.04 c ]]> TR 421±10 <![CDATA[2398±40 a ]]> <![CDATA[94.1±1.6 a ]]> <![CDATA[1.63±0.03 a ]]> Note: Different letters in the superscript of the same data indicate significant differences (P<0.05), while the same letter indicates no significant differences (P>0.05).

[0053] 2. Gut health and disease resistance like Figure 6 As shown, on day 4 after challenge (day 25 of the experiment), broilers in the challenge control group (PC) showed typical intestinal damage, with their jejunal lesion scores significantly higher than those of other groups (P<0.05). The lesion scores of the treatment group (TR) were significantly lower than those of the PC group (P<0.01), and there was no significant difference compared to the NC and NS groups.

[0054] Reference Figure 6 , Figure 7 In the count of Clostridium perfringens in cecal chyme, the colony count in the PC group (7.2 ± 0.3 lg CFU / g) was significantly higher than that in the NC group (4.1 ± 0.2 lg CFU / g) and the NS group (3.9 ± 0.2 lg CFU / g). Although the Clostridium perfringens count in the TR group (5.0 ± 0.3 lg CFU / g) was higher than that in the non-challenge group, it was significantly lower than that in the PC group (P<0.05), indicating that the product of this invention can effectively inhibit the proliferation of pathogenic bacteria in the intestine.

[0055] in conclusion: This embodiment fully demonstrates that by adding 0.1% of the anti-corrosion nano-selenium compound liquid of the present invention to drinking water, it is possible to: It significantly promotes the growth of broiler chickens, increases daily weight gain, and reduces feed conversion ratio; It effectively alleviates the negative effects of combined coccidia and Clostridium perfringens infection, significantly reduces the severity of intestinal lesions and the number of pathogens, and demonstrates good preventive and therapeutic effects; It achieves the dual functions of selenium nutritional supplementation and intestinal health regulation. Its mechanism of action may be closely related to the synergistic antioxidant effects of nano-selenium, active metabolites of Bacillus licheniformis (antimicrobial peptides and digestive enzymes), and vitamins E and C, which are abundant in the product.

[0056] This invention utilizes the fermentation supernatant of Bacillus licheniformis B1-Se under specific conditions to completely replace pure water as the solvent and medium for the chemical synthesis reaction of nano-selenium. This is not a simple solvent replacement, but rather the introduction of a biologically active, functionalized, and complex system. The supernatant itself is rich in antibacterial metabolites (such as Surfactin and Fengycin), digestive enzymes, residual sodium selenite, and live bacteria. As a "functionalized matrix," it fundamentally reconstructs the microecological environment of traditional nano-selenium liquids, giving them inherent anti-putrefaction and antioxidant capabilities. Furthermore, it exhibits several synergistic effects in animal waterline feeding applications: ① Synergistic antibacterial effect: The natural antimicrobial peptides in the supernatant and the stable acidic environment (pH 4.0) modulated by citric acid together form a powerful antimicrobial microenvironment.

[0057] ② Synergistic antioxidant effect: The bacterial metabolic antioxidants, vitamin C (reducing agent) and vitamin E in the supernatant together form a multi-layered antioxidant network, which effectively protects the nano-selenium and prevents it from oxidizing and turning black.

[0058] ③ Synergistic Functions: The final product integrates "nutrition (nano-selenium supplementation) + health care (antibacterial, antioxidant, digestive aid) + stability (anti-corrosion, anti-blackening)," surpassing the scope of traditional single-function selenium supplements.

[0059] In the description of this specification, the references to terms such as "one embodiment," "some embodiments," "example," "specific example," or "some examples," etc., indicate that a specific feature, structure, material, or characteristic described in connection with that embodiment or example is included in at least one embodiment or example of the present invention. In this specification, the illustrative expressions of the above terms do not necessarily refer to the same embodiment or example. Furthermore, the specific features, structures, materials, or characteristics described may be combined in any suitable manner in one or more embodiments or examples. Moreover, without contradiction, those skilled in the art can combine and integrate the different embodiments or examples described in this specification, as well as the features of different embodiments or examples.

[0060] The above description is merely a preferred embodiment of the present invention and is not intended to limit the present invention in any way. Although the present invention has been disclosed above with reference to preferred embodiments, it is not intended to limit the present invention. Any person skilled in the art can make some modifications or alterations to the above-described technical content to create equivalent embodiments without departing from the scope of the present invention. The implementation schemes in the above embodiments can also be further combined or replaced. Any simple modifications, equivalent changes and alterations made to the above embodiments based on the technical essence of the present invention without departing from the scope of the present invention shall still fall within the scope of the present invention.

Claims

1. A method for preparing a nano-selenium compound liquid, characterized in that, Includes the following steps: S1. Provide Bacillus licheniformis strain B1-Se, wherein the preservation number of Bacillus licheniformis strain B1-Se is GDMCCNO:64806; S2. Fermentation is carried out using the strain in a culture medium containing a carbon source, a nitrogen source, and sodium selenite; S3. After fermentation, the fermentation broth is separated into solid and liquid components, and the supernatant is collected. S4. Using the supernatant obtained in step S3 as a solvent, adjust its pH to 3.8-4.2, and then add dispersant, ascorbic acid and sodium selenite in sequence to react and generate nano-selenium liquid. S5. Add vitamin E to the nano-selenium liquid obtained in step S4, mix well, and obtain the compound liquid.

2. The method as described in claim 1, characterized in that, In step S2: The carbon source is sucrose with a concentration of 25 g / L; The nitrogen source is ammonium sulfate with a concentration of 10 g / L; The concentration of sodium selenite added was 1.1 g / L.

3. The method as described in claim 1, characterized in that: In step S4, citric acid is used to adjust the pH.

4. The method as described in claim 1, characterized in that: In step S4, adjust the pH to 4.

0.

5. The method according to claim 1, characterized in that, In step S4: The dispersant is chitosan, and the addition amount is 0.0975 kg / t; The amount of ascorbic acid added is 8.25 kg / t; The amount of sodium selenite added resulted in a total selenium content of 0.2% w / v in the final product.

6. The method as described in claim 1, characterized in that, In step S5: The vitamin E mentioned is modified water-soluble vitamin E with an effective content of 50%, and its concentration added to the final product is 10 kg / t.

7. A nano-selenium complex liquid prepared by the method according to any one of claims 1-6, characterized in that: The nano-selenium compound liquid contains nano-elemental selenium, fermentation metabolites of Bacillus licheniformis B1-Se, dispersant, ascorbic acid, and vitamin E.

8. The nano-selenium compound liquid as described in claim 7, characterized in that: The content of the nano-elemental selenium is 0.2% w / v; The fermentation metabolites of Bacillus licheniformis B1-Se include antimicrobial lipopeptides and digestive enzymes.

9. The nano-selenium compound liquid as described in claim 7, characterized in that: The nano-selenium compound liquid also contains live Bacillus licheniformis B1-Se bacteria, with a live bacteria count of not less than 1×10⁻⁶. 6 cfu / mL.

10. The application of the nano-selenium compound liquid as described in any one of claims 7-9 in the preparation of animal drinking water additives.

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