Biosynthesis-based dry grass feed for fragrant oil palm and preparation method thereof
By integrating Weissella fermentation of Cyperus rotundus with Cyperus rotundus meal water extract and urea, the nutritional structure and flavor of Cyperus rotundus feed were improved, solving the problems of unutilized nutrients and weakened aroma in existing technologies, and achieving efficient fermentation and flavor enhancement of the feed.
Patent Information
- Application Number
- CN202311373618.8
- Authority / Receiving Office
- CN · China
- Patent Type
- Patents(China)
- Current Assignee / Owner
- Filing Date
- 2023-10-23
- Publication Date
- 2026-02-24
- Estimated Expiration
- 2043-10-23
AI Technical Summary
In existing technologies, the nutrients in the stems and leaves of Cyperus rotundus are not fully utilized, and its unique aroma components are weakened during processing. Microbial fermentation technology has failed to effectively improve the nutritional structure and flavor of the straw.
Weissella confusa was used to ferment Cyperus rotundus, combined with Cyperus rotundus meal extract and urea as carbon and nitrogen sources. Through fermentation, the dried Cyperus rotundus produced beneficial volatile substances, which improved the aroma and nutritional composition of the feed.
It improves the aroma and nutritional value of feed, inhibits mold growth, reduces nutrient loss, and enhances palatability and utilization of feed.
Smart Images
Figure CN117752012B_ABST
Abstract
Description
Technical Field
[0001] This invention belongs to the field of feed fermentation, specifically relating to a strongly aromatic tiger nut hay feed based on biosynthesis and its preparation method. Background Technology
[0002] Crop straw already plays a significant role in the animal feed industry. However, feeding animals directly with untreated straw results in incomplete absorption of nutrients, failing to fully realize its nutritional value. Microbial fermentation of straw breaks down cellulose and hemicellulose, improving nutrients, enhancing the original quality and structure of the straw, and generating beneficial metabolic small molecules, thus greatly increasing the utilization rate and feed value of straw. Therefore, this invention utilizes *Westernella* to ferment *Cyperus rotundus* to prepare a multi-nutritional and flavorful hay feed.
[0003] Studies have shown that the above-ground stems and leaves of tiger nuts are nutritionally complete and rich, containing nutrients such as crude fat, protein, sugar, and crude fiber. At present, tiger nuts are generally processed directly into hay (i.e., tiger nuts hay), and the nutrients are not fully utilized, while the unique aroma components of tiger nuts are weakened.
[0004] The health effects of *Westernella fusionis* on humans and livestock may suggest its potential as a probiotic. It can colonize the gut, overcoming the acidic conditions of the stomach and the bile salt environment, thereby exerting beneficial effects such as regulating gut microbiota balance and activating the host's endogenous microbiota or immune activity. Fermentation of *Westernella fusionis* produces short-chain fatty acids and alcohols, which, through technical processing, can undergo esterification to generate volatile esters. Summary of the Invention
[0005] The purpose of this invention is to overcome the shortcomings of the prior art and provide a strongly aromatic tiger nut hay feed based on biosynthesis and its preparation method.
[0006] To achieve the above objectives, the technical solution adopted by the present invention is as follows:
[0007] A biosynthesized aromatic oilseed hay feed includes oilseed hay, a bacterial solution of *Weissella fusionis*, and carbon and nitrogen sources; the *Weissella fusionis* has been deposited at the China General Microbiological Culture Collection Center (CGMCC) with accession number CGMCC No. 24053.
[0008] The carbon source is an aqueous extract of tiger nut meal, and the nitrogen source is urea.
[0009] The fused Weissl bacterial suspension is prepared by the following method: after high-pressure sterilization of MRS medium, Weissl fusion bacteria are inoculated and activated to obtain the fused Weissl bacterial suspension.
[0010] The activation and expansion conditions are as follows: Activation conditions: After thawing the bacterial culture stored at -80℃, inoculate it into sterilized MRS medium at a 3% inoculum rate and incubate at 37℃ and 180 r / min for 6-8 hours; Expansion conditions: Inoculate the activated bacterial culture into MRS medium at a 2% inoculum rate and incubate under the same conditions for 6-8 hours to achieve a bacterial concentration of 10. 8 CFU / mL.
[0011] The amount of the fused Weissella bacterial solution added is 2 ml / 100g relative to the oilseed hay.
[0012] The aqueous extract of tiger nut meal was prepared as follows: tiger nut meal and distilled water were added and mixed at a mass ratio of 1:8. The mixture was then extracted by magnetic stirring in a constant temperature water bath at 50°C for 1 hour, centrifuged at 3000 r / min for 10 minutes, and the supernatant was taken as the aqueous extract of tiger nut meal.
[0013] The present invention also includes a method for preparing the aromatic oilseed hay feed based on biosynthesis, comprising the following steps: adding a bacterial solution of fused Weissella bacteria, a nitrogen source and a carbon source to the oilseed hay, mixing them evenly, spraying a small amount of distilled water evenly onto the straw, adjusting the moisture content, and fermenting to obtain the product.
[0014] The moisture content was adjusted to 65%.
[0015] The fermentation conditions were as follows: the contents were compacted, vacuum-packed in fermentation bags, and then placed in a 35 ℃ constant temperature incubator for 7 days.
[0016] The oilseed sedge is cut to 1-1.5cm.
[0017] Compared with the prior art, the beneficial effects of the present invention are:
[0018] The technical solution of this application utilizes biotechnology to assist in the fermentation of Cyperus rotundus. By decomposing fibers and utilizing soluble carbohydrates and other fermentation substrates to produce organic acids, it inhibits the growth of molds and other putrefactive bacteria, reducing the loss of nutrients. At the same time, the flavor substances (mainly including alcohols, aldehydes, phenols, acids, esters, ketones and their hydrocarbons) are greatly enriched after fermentation, which can effectively increase the aroma components of the feed.
[0019] Cyperus rotundus is rich in cellulose and hemicellulose. Cellulose is a high-molecular-weight polymer composed of glucose linked by β-1,4 glycosidic bonds. Its linear chains are parallel to each other, do not exhibit a helical conformation, and have no branching structure. The intra- and inter-chain hydrogen bonds stably bind the molecules, forming a water-insoluble polymer with a unique three-dimensional network structure and porous adsorption capacity. However, the cellulose in Cyperus rotundus hay contains hemicellulose, lignin, and other substances, causing the molecules to be tightly bound together, forming highly ordered crystalline regions, and the porous material is not exposed. This application modifies the substrate through fusion fermentation with *Westernella*. Figure 1 (The schematic diagram is shown). At the same time, proteases and cellulases are generated, which break or form non-covalent bonds such as hydrogen bonds and ionic bonds in the attached material and the treated material. After vibration, the porous structure of cellulose is exposed, and the aromatic substances produced are adsorbed, thereby effectively increasing the aroma components of the feed. Attached Figure Description
[0020] Figure 1 Schematic diagram illustrating the principle of porous cellulose adsorption of volatile substances;
[0021] Figure 2 An appearance diagram of Cyperus rotundus and the Cyperus rotundus feed of this application;
[0022] Figure 3 Microscopic structures of dried oilseed grass before and after fermentation;
[0023] Figure 4 FTIR spectra of dried oilseed grass before and after fermentation;
[0024] Figure 5 Heat maps of flavor compounds in dried sedge before and after fermentation;
[0025] Figure 6 This is a graph showing the changes in the types and relative contents of volatile flavor compounds in dried oilseed cake before and after fermentation. Detailed Implementation
[0026] To enable those skilled in the art to better understand the technical solution of the present invention, the present invention will be further described in detail below with reference to the accompanying drawings and preferred embodiments.
[0027] This invention relates to *Westernella fusionis* isolated from fresh human feces, deposited at the China General Microbiological Culture Collection Center (CGMCC) with accession number CGMCC No. 24053, deposited on October 8, 2021, at No. 3, Courtyard 1, Beichen West Road, Chaoyang District, Beijing. The proposed classification and nomenclature for the deposited biological material is *Westernella fusionis* (…). Weissella confusa ). Example
[0028] The preparation method of aromatic oilseed hay feed based on biosynthesis adopts the following steps:
[0029] 1) Preparation of bacterial suspension from *Westernella*
[0030] After thawing the bacterial culture stored at -80℃, inoculate it into sterilized MRS medium at a 3% inoculum rate and incubate at 37℃ and 180 r / min for 6-8 hours. For expansion, inoculate the activated bacterial culture into MRS medium at a 2% inoculum rate and incubate under the same conditions for 6-8 hours to achieve a bacterial concentration of 10⁻⁶. 8 CFU / mL.
[0031] 2) Preparation of aqueous extract of tiger nut meal
[0032] Add tiger nut meal and distilled water in a ratio of 1:8, mix well, and extract with magnetic stirring in a 50℃ constant temperature water bath for 1 hour. Centrifuge at 3000 r / min for 10 minutes, and take the supernatant as the desired water extract.
[0033] 3) Feed preparation
[0034] Cut the dried stalks of Cyperus rotundus to about 1-1.5 cm. Weigh 100 g and add the following to the stalks: a bacterial solution of Bacillus fragilis (2 mL / 100 g dried stalks), a water extract of Cyperus rotundus soybean meal (6 g / 100 g stalks), and urea (0.5 g / 100 g stalks).
[0035] After mixing evenly, a small amount of distilled water is sprayed evenly onto the straw to adjust the moisture content to 65%. The straw is then compacted, packed into fermentation bags, vacuum-packed, and placed in a 35℃ constant temperature incubator for fermentation for 7 days.
[0036] 1) Product Form Display
[0037] like Figure 2 As shown in (a), unfermented sedge is greenish in color, has a fragrant aroma, abundant leaves, and soft stems; after fermentation, the material becomes softer. Figure 2 (b) ), with a rich grassy aroma.
[0038] 2) Impact on fermentation quality
[0039] To better illustrate the beneficial effects of adding Fusobacterium fusion, tiger pea meal water extract, urea, etc., this experiment selected different additive types as control examples to prepare different feeds. The treatment groups are shown in Table 1.
[0040] Table 1
[0041] 2.1 Feed Nutritional Components
[0042] ①Basic components
[0043] Table 2 shows that anaerobic fermentation increased the content of crude protein, fat, and dry matter in straw in this experiment. Taking crude protein as an example, the crude protein content of group WC-4 increased by 43.83% after fermentation (compared to CK). This indicates that adding tiger nut meal aqueous extract and urea can provide microorganisms with sufficient carbon and nitrogen sources, thereby slowing down the decomposition of carbohydrates in straw itself.
[0044] The content of neutral detergent fiber and acid detergent fiber in feed is related to animal feed intake and digestibility, and is negatively correlated with the quality of straw fermentation. WC-4 has the lowest content of neutral detergent fiber and acid detergent fiber. Table 2 shows the nutrient composition of Cyperus rotundus straw before and after fermentation (unit: %).
[0045] Table 2
[0046] ② Organic acid content
[0047] The content of organic acids is another important indicator for evaluating the quality of fermented products. The production of organic acids acidifies the feed, quickly creating a low-acid environment, lowering the pH value, and inhibiting the growth of aerobic microorganisms, thus effectively maintaining its quality. As shown in Table 3, the contents of lactic acid, acetic acid, and propionic acid in each treatment group were higher than those in the control group. *Westernella*, as a heterologous lactic acid bacterium, can produce byproducts such as acetic acid, which inhibits the reproduction of harmful bacteria, in addition to lactic acid, during the fermentation process, thereby improving the fermentation success rate. Table 3 shows the organic acid content of fermented straw (unit: mg / mL).
[0048] Table 3
[0049] 2.2 Changes in feed structure
[0050] ① Microstructure observation
[0051] The microstructure of the feed was observed, and the results were as follows: Figure 3 Significant changes were observed in the microstructure of the samples before and after treatment. In the crude sedge, the pores on the surface were closed, surrounded by numerous fine particles. The pores and their appendages on the epidermis were arranged in a regular pattern. The surface structure of the sample with only distilled water added (CK) was not damaged, the pores were opened, and the surface fibers were partially utilized. The surfaces of the other treatment groups showed irregular changes, with open pores, grooves appearing on the fiber surface, and fewer surrounding particles. As fermentation progressed, the fibers were utilized and degraded by microorganisms, and the surface became loose and cracked.
[0052] ②Fourier Infrared Spectroscopy Analysis
[0053] The chemical structure of Cyperus rotundus straw before and after fermentation was observed by Fourier transform infrared spectroscopy (FTIR). Figure 4 As shown, similar characteristic absorption peaks were observed in all samples, with slight variations in peak intensity at specific wavenumbers, specifically at 3415.90 cm⁻¹. -1 2921.00 cm-1 1617.38 cm -1 1046.45 cm -1 A characteristic absorption peak appears near the same wavelength. (3400-3430 cm⁻¹) -1 The absorption band at 2900-2700 cm⁻¹ is likely due to the stretching of OH groups in cellulose. -1 The weaker absorption band between them may be due to the reverse stretching vibration of -CH in carbohydrates and lignin, 1617.38 cm⁻¹ -1 The location is attributed to the stretching of cellulose and lignin C=O, 1046.45 cm. -1 This is due to CC stretching. It indicates that the cellulose and hemicellulose in the fermented oilseed rape straw have degraded, and the microscopic surface structure of the straw has been altered.
[0054] 2.3 Determination of Volatile Matter in Feed
[0055] The changes in volatile flavor compounds of Cyperus rotundus before and after fermentation were analyzed using GC-MS. A visualization heatmap of the relative contents of the detected volatile flavor compounds was plotted (e.g., Figure 5 As shown in the figure, the darker the color, the higher the relative content. A total of 188 flavor compounds were detected in the treatment groups, including 13 aldehydes, 26 hydrocarbons, 5 phenols, 29 esters, 32 ketones, 8 terpenes, 66 alcohols, 5 acids and 4 other substances.
[0056] The flavor compounds mainly include alcohols, aldehydes, phenols, acids, esters, ketones, and hydrocarbons. Fermentation not only increases the variety of flavor compounds in the feed but also imparts more aroma types, making the aroma more intense. Compared with the CK0 group, the CK, WC-2, and WC-4 groups showed a decrease of 11, 2, 5, and 7 flavor compounds, respectively, while the remaining treatment groups showed an increase in flavor compounds. After straw fermentation, the relative content of aldehydes decreased, mainly providing a fruity aroma. The relative content of phenols increased compared to the CK0 group, with WC-4 increasing from 0.36% to 56.61%. The relative content of flavonoids increased in the treatment groups, while it decreased in the CK control group. Phenolic and flavonoid substances can effectively enhance the antioxidant properties of the samples, thus contributing to the stability of sample quality. Furthermore, studies have found that microbial fermentation significantly reduces or even completely degrades the content of toxic substances in straw, increasing flavor and imparting a special aroma, reducing the toxicity of the raw material, and improving its palatability and utilization rate.
[0057] Based on the above experiments, sedge hay is rich in cellulose and hemicellulose. Cellulose is a high-molecular-weight polymer composed of glucose linked by β-1,4 glycosidic bonds. Its straight chains are parallel to each other, do not exhibit a helical conformation, and have no branching structure. The intra- and inter-chain hydrogen bonds stably bind the molecules, forming a water-insoluble polymer with a unique three-dimensional network structure and porous adsorption capacity. Meanwhile, the cellulose in hay is mixed with hemicellulose, lignin, and other substances, causing the molecules to be tightly bound together, forming highly ordered crystalline regions, and the porous materials are not exposed.
[0058] Fusion fermentation with *Westernella* modifies the substrate, simultaneously producing proteases and cellulases. These proteases and cellulases disrupt or degrade non-covalent bonds such as hydrogen bonds and ionic bonds in the treated material, exposing the porous structure of cellulose. Figure 5 It can be verified that the cellulose and hemicellulose in the sedge were degraded after fermentation, and Figure 3 This provides a more direct view of the open pores in the feed after fermentation. Furthermore, GC-MS analysis... Figure 5 and Figure 6 Feed produces more volatile substances. Figure 6 Types of volatile flavor compounds in sedge straw before and after fermentation ( Figure 6 (a) and relative content changes ( Figure 6 (b)), its principle is as follows Figure 1 As shown, Figure 1 (a) shows the fiber structure of dried sedge, with cellulose, hemicellulose, lignin and other components cross-linked; (b) after modification with Fussella fusion, the degree of cross-linking is reduced, revealing a porous three-dimensional structure; (c) the treated material shows a porous structure that can adsorb the generated aromatic substances and retain the aroma components.
[0059] In summary, the technical solution of this application utilizes biotechnology to assist in the fermentation of Cyperus rotundus. By decomposing fibers and utilizing soluble carbohydrates and other fermentation substrates to produce organic acids, it inhibits the growth of molds and other putrefactive bacteria, reducing nutrient loss. Simultaneously, the fermentation process greatly enriches flavor compounds (mainly including alcohols, aldehydes, phenols, acids, esters, ketones, and hydrocarbons), effectively increasing the aroma components of the feed.
[0060] The above description is merely a preferred embodiment of the present invention. It should be noted that those skilled in the art can make various improvements and modifications without departing from the principles of the present invention, and these improvements and modifications should also be considered within the scope of protection of the present invention.
Claims
1. A fusion of Weissella bacteria, characterized in that, The aforementioned fusion Weissella is Weissella confusa ZZK, which has been deposited at the China General Microbiological Culture Collection Center (CGMCC) with accession number CGMCC No. 24053 and deposit date of December 8, 2021.
2. An application of the fused Weissella strain as described in claim 1, characterized in that, This invention relates to a biosynthesized aromatic oilseed hay feed, which is prepared by the following steps: adding a bacterial solution of *Westernella* fused with oilseed hay, a nitrogen source, and a carbon source to oilseed hay, mixing them evenly, spraying a small amount of distilled water evenly onto the straw, adjusting the moisture content, and fermenting to obtain the final product.
3. A method for preparing a fragrant oilseed hay feed based on biosynthesis, characterized in that, The following steps were adopted: A bacterial solution of *Weissella confusa*, a nitrogen source, and a carbon source were added to the dried oilseed straw. After mixing evenly, a small amount of distilled water was mixed evenly and sprayed onto the straw to adjust the moisture content. Fermentation was then carried out. The *Weissella confusa* strain was identified as *Weissella confusa* ZZK, which was deposited at the China General Microbiological Culture Collection Center (CGMCC) with accession number CGMCC No. 24053 and a deposit date of December 8, 2021.
4. The application of the biosynthesized aromatic sedge hay feed as described in claim 2, or the preparation method of the biosynthesized aromatic sedge hay feed as described in claim 3, characterized in that, The carbon source is an aqueous extract of tiger nut meal, and the nitrogen source is urea.
5. The application of the biosynthesized aromatic sedge hay feed as described in claim 2, or the preparation method of the biosynthesized aromatic sedge hay feed as described in claim 3, characterized in that, The amount of the fused Weissella bacterial solution added is 2 mL / 100g relative to the oilseed hay.
6. The application of the biosynthesized aromatic sedge hay feed as described in claim 2, or the preparation method of the biosynthesized aromatic sedge hay feed as described in claim 3, characterized in that, The aqueous extract of tiger nut meal was prepared as follows: tiger nut meal and distilled water were added and mixed at a mass ratio of 1:
8. The mixture was then extracted by magnetic stirring in a constant temperature water bath at 50°C for 1 hour, centrifuged at 3000 r / min for 10 minutes, and the supernatant was taken as the aqueous extract of tiger nut meal.
7. The application of the biosynthesized aromatic sedge hay feed as described in claim 2, or the preparation method of the biosynthesized aromatic sedge hay feed as described in claim 3, characterized in that, The moisture content was adjusted to 65%.
8. The application of the biosynthesized aromatic sedge hay feed as described in claim 2, or the preparation method of the biosynthesized aromatic sedge hay feed as described in claim 3, characterized in that, The fermentation conditions were as follows: the contents were compacted, vacuum-packed in fermentation bags, and then placed in a 35 ℃ constant temperature incubator for 7 days.
9. The application of the biosynthesized aromatic oilseed hay feed as described in claim 2, or the preparation method of the biosynthesized aromatic oilseed hay feed as described in claim 3, characterized in that, The oilseed sedge is cut to 1-1.5cm.
10. The biosynthetic aromatic oilseed hay feed obtained by the preparation method of the biosynthetic aromatic oilseed hay feed according to any one of claims 3-9.