CN117694461B - Application of angelic acid and/or 4-cyclopentene-1, 3-dione in preparation of choline degradation inhibitor - Google Patents
Application of angelic acid and/or 4-cyclopentene-1, 3-dione in preparation of choline degradation inhibitor Download PDFInfo
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- CN117694461B CN117694461B CN202410035631.0A CN202410035631A CN117694461B CN 117694461 B CN117694461 B CN 117694461B CN 202410035631 A CN202410035631 A CN 202410035631A CN 117694461 B CN117694461 B CN 117694461B
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- A—HUMAN NECESSITIES
- A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
- A23K—FODDER
- A23K50/00—Feeding-stuffs specially adapted for particular animals
- A23K50/10—Feeding-stuffs specially adapted for particular animals for ruminants
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- A—HUMAN NECESSITIES
- A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
- A23K—FODDER
- A23K20/00—Accessory food factors for animal feeding-stuffs
- A23K20/10—Organic substances
- A23K20/105—Aliphatic or alicyclic compounds
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- A—HUMAN NECESSITIES
- A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
- A23K—FODDER
- A23K50/00—Feeding-stuffs specially adapted for particular animals
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- A—HUMAN NECESSITIES
- A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
- A23K—FODDER
- A23K50/00—Feeding-stuffs specially adapted for particular animals
- A23K50/70—Feeding-stuffs specially adapted for particular animals for birds
- A23K50/75—Feeding-stuffs specially adapted for particular animals for birds for poultry
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Abstract
本发明公开了当归酸和/或4‑环戊烯‑1,3‑二酮在制备胆碱降解抑制剂中的应用。本发明首次发现当归酸和4‑环戊烯‑1,3‑二酮可抑制胃肠道微生物对胆碱的降解，作为饲料添加剂在抑制胆碱降解的同时，4‑环戊烯‑1,3‑二酮还可提高饲料利用率。The present invention discloses the use of angelic acid and/or 4-cyclopentene-1,3-dione in the preparation of a choline degradation inhibitor. The present invention firstly finds that angelic acid and 4-cyclopentene-1,3-dione can inhibit the degradation of choline by gastrointestinal microorganisms. As a feed additive, 4-cyclopentene-1,3-dione can inhibit the degradation of choline and improve feed utilization.
Description
Technical Field
The invention relates to application of angelic acid and/or 4-cyclopentene-1, 3-dione in inhibiting gastrointestinal tract microorganism from degrading choline, belonging to the field of compound application.
Background
Choline is an essential nutrient for animals and can affect various physiological processes of the liver, such as lipid and cholesterol metabolism. Although the liver can produce some choline, choline is mainly obtained from feed. Studies have shown that reduced choline levels induce fatty liver disease by reducing the efflux of very low density lipoproteins from hepatocytes. Choline is critical to perinatal cows. Perinatal cows generally have reduced incidence of fatty liver and ketosis by feeding ruminally protected choline chloride. However, gastrointestinal microorganisms rapidly degrade choline in feed to trimethylamine, resulting in reduced bioavailability of choline by animals, which may be associated with fatty liver or ketosis in some of the cows. In addition, trimethylamine and its metabolite in liver oxidize trimethylamine to have toxic effect on animals and harm animal health. Trimethylamine is also an important substance that causes animal products to develop fishy smell. However, there is currently no choline degradation inhibitor suitable for animal use.
Disclosure of Invention
The first object of the invention is to provide the application of angelic acid and/or 4-cyclopentene-1, 3-dione in preparing choline degradation inhibitors. The second object of the present invention is to provide the use of angelic acid and/or 4-cyclopentene-1, 3-dione in the preparation of feed additives.
The technical scheme is that the application of angelic acid and/or 4-cyclopentene-1, 3-dione in preparing choline degradation inhibitors.
The choline degradation inhibitor comprises angelic acid and/or 4-cyclopentene-1, 3-dione. The invention relates to an application of angelic acid and/or 4-cyclopentene-1, 3-dione in preparing a feed additive, wherein the feed additive is used for inhibiting degradation of choline in gastrointestinal tracts.
Wherein the feed additive is used for reducing the content of trimethylamine oxide and trimethylamine in livestock blood.
Wherein the livestock comprises one of ruminants, monogastric animals and birds.
The active ingredients of the feed additive are angelic acid and/or 4-cyclopentene-1, 3-dione.
Wherein the feed additive is used for reducing fishy smell of livestock products caused by trimethylamine.
Wherein the 4-cyclopentene-1, 3-dione is used for improving the utilization rate of the feed.
Compared with the prior art, the invention has the remarkable advantages that the invention discovers that the angelic acid and the 4-cyclopentene-1, 3-dione can inhibit the degradation of choline by gastrointestinal microorganisms for the first time, and can also improve the utilization rate of the feed when being used as a feed additive to inhibit the degradation of choline in animal gastrointestinal tracts.
Detailed Description
The technical scheme of the invention is further described below with reference to the specific technical scheme.
Example 1
1. Materials and methods
1.1 Strain and Medium and preparation
1.1.1 Strain
The choline degrading bacteria used in this study were ESCHERICHIA FERGUSONII ATCC 35469.
1.1.2 Medium and configuration
The medium composition is shown in Table 1.
TABLE 1 Medium composition (1.0L)
The preparation method of the reagent comprises the following steps:
The preparation of the gastric juice without cytoma comprises filtering rumen juice with two layers of gauze to remove large granule residues, centrifuging filtrate at 12,000rpm/min at 4deg.C for 20min, collecting supernatant, and storing at-20deg.C for use.
The microelement solution is prepared by dissolving nitrilotriacetic acid (nitrilotriacetic acid)(1.5g)、NaCl(1.0g)、NaMoO4·2H2O(0.01g)、NaSeO4(0.19g)、Na2WO4·2H2O(0.1g)、MgSO4·7H2O(3.0g)、AlK(SO4)2·12H2O(0.018g)、CoSO4·7H2O(0.18g)、CaCl2·2H2O(0.1g)、ZnSO4·7H2O(0.18g)、FeSO4·7H2O(0.1g)、MnSO4.H2O(0.45g)、NiSO4·6H2O(0.1g)、CuSO4·5H2O(0.01g)、H3BO3(0.01g) in 1L deionized water.
The preparation of fatty acid solution comprises the steps of weighing 8.0g of NaOH, dissolving in 800mL of deionized water, adding 6.85mL of acetic acid, 3.00mL of propionic acid, 1.84mL of butyric acid and 0.55mL of valeric acid, adding a proper amount of deionized water to a constant volume to 1L, and preserving at 4 ℃ for later use.
The preparation of vitamin solution comprises weighing 54.7g KH 2PO4, dissolving in 1L deionized water, sequentially adding 20.4mg biotin, 20.4mg folic acid, 164mg calcium pantothenate, 164mg nicotinamide, 164mg riboflavin, 164mg thiamine hydrochloride, 164mg vitamin B6 hydrochloride, 20.4mg para-aminobenzoic acid and 20.5mg cobalamin, filtering, sterilizing, packaging in anaerobic tube, and preserving at 4deg.C for use.
1.1.3 Preparation of Medium
The preparation (1L) of anaerobic culture medium comprises accurately measuring each component in wide-bottom flask according to the culture medium formula, adding deionized water to make up 1L, heating and boiling in a microwave oven until the liquid changes from blue to red, rapidly taking out, immediately introducing carbon dioxide for about 2h, then adding L-cysteine hydrochloride until the liquid becomes clear, packaging, introducing carbon dioxide in advance to remove air in a fermentation tube, adding 9.6mL of culture medium, sealing the bottle mouth with a rubber plug, sealing with an aluminum cap, placing into an autoclave for sterilization, and cooling for later use.
1.2 Design of experiments
Angelic acid solution or 4-cyclopentene-1, 3-dione was added to the fermenter respectively in the amounts shown in Table 2,4 replicates per group. 20.0mmol/L choline is added into each fermentation tube as a substrate, and the bacterial liquid which is cultivated until the logarithmic phase is inoculated into a culture medium which is injected with 0.1mL of vitamin solution and angelic acid solution/4-cyclopentene-1, 3-dione in advance, wherein the inoculation amount is 3.0 percent, and the bacterial liquid is cultivated for 60 hours at 39 ℃ and 50 rpm/min.
The amount of inhibitor added (final concentration) in Table 2
1.3 Sample collection and index determination
1.3.1 Growth Curve drawing
At the end of fermentation, the absorbance (OD value) of the bacteria liquid in the fermentation tube after shaking was measured by a spectrophotometer at a wavelength of 600 nm.
1.3.2 Choline concentration determination
1.0ML of the fermentation broth was centrifuged at 13000 Xg at 4℃for 2min, and the supernatant was collected. Concentration was measured using a bovine choline ELISA assay kit (Emblica Corp., china) and the test procedure was described in the specification.
1.4 Statistical processing of data
Data were analyzed using One-way ANOVA (Tukey) in SPSS20.0 software, P <0.05 indicated significant differences.
2. Results
As shown in tables 3 and 4, angelic acid and 4-cyclopentene-1, 3-dione can inhibit ESCHERICHIA FERGUSONII degradation of choline in a targeting manner, and can inhibit choline degradation under certain concentration conditions, but basically does not influence microbial growth.
TABLE 3 Effect of angelic acid on ESCHERICHIA FERGUSONII degradation of choline
Note that the shoulder marks of the same row of data show insignificant differences (P > 0.05), the different letters show significant differences (P < 0.05), the initial concentration of choline is 20.0mmol/L, and the four concentrations of angelic acid have no effect on the degradation of choline without adding ESCHERICHIA FERGUSONII (19.6, 19.8, 19.9, 19.5 mmol/L).
TABLE 4 influence of 4-cyclopentene-1, 3-dione on ESCHERICHIA FERGUSONII degradation of choline
Note that the shoulder marks of the same row of data show insignificant differences (P > 0.05), the different letters show significant differences (P < 0.05), the initial concentration of choline is 20.0mmol/L, and 4-cyclopentene-1, 3-dione at four concentrations has no effect on the degradation of choline without adding ESCHERICHIA FERGUSONII (19.5, 19.8, 19.7, 19.5 mmol/L).
Example 2
1 Material method
1.1 Experimental materials
1.1.1 Test animals and rumen fluid collection
The test animals were raised at the university of Nanjing agriculture animal science and technology college dairy cow test base (Nanjing, jiangsu), 3 Holstein cows with permanent rumen fistula had an average body weight of 613+ -35 kg. On the day of the test, rumen fluid was taken half an hour before morning feeding and quickly returned to the laboratory in a vacuum flask that had been pre-filled with carbon dioxide.
1.1.2 Preparation of Artificial rumen fermentation broth
The collected fresh rumen fluid is filtered by 4 layers of gauze, and the rumen fluid and buffer solution (specific components are shown in table 5) are mixed according to the proportion of 1:9 (v/v) to prepare the mixed artificial rumen culture fluid. The mixed fermentation liquor is packaged into 180mL fermentation bottles, the temperature in the whole process is kept at 39 ℃, pure carbon dioxide is continuously introduced to keep anaerobic environment, and a magnetic stirrer is used for stirring to keep the rumen liquor and the buffer solution uniformly mixed. 1g of straw powder and inhibitor (angelic acid or 4-cyclopentene-1, 3-dione) are added into a fermentation bottle in advance, and are placed into a constant temperature incubator at 39 ℃ for preheating for 30min. After the completion of the split charging, the culture was carried out at 39℃and 80r/min for 72 hours in a constant temperature shaker.
TABLE 5 buffer composition (1L)
Preparing a specific solution:
(1) Microelement solution (A liquid)
Deionized water was added to 100mL
(2) Buffer solution (liquid B)
NH4HCO3 4.0g
NaHCO3 35.0g
Deionized water was added to 1000mL
(3) Macroelement solution (C liquid)
Na2HPO4·12H2O 9.45g
KH2PO4 6.2g
MgSO4·7H2O 0.6g
Deionized water was added to 1000mL
(4) 0.1% Resazurin solution (solution D)
100Mg of resazurin was added to 100mL of deionized water.
(5) Reducing agent solution (E liquid)
1.2 Test methods
1.2.1 Test design
Example 1 test set 4 groups.
1. Treatment group 1, no drug added.
2. Treatment group 2 angelic acid 1.8mmol/L.
3. Treatment group 3 angelic acid 3.6mmol/L.
4. Treatment group 4 angelic acid 7.2mmol/L.
5. Treatment group 5 4-cyclopentene-1, 3-dione 1.8mmol/L.
6. Treatment group 6 4-cyclopentene-1, 3-dione 3.6mmol/L.
7. Treatment group 7:4-cyclopentene-1, 3-dione 7.2mmol/L
1.2.2 Chemical analysis
And (3) measuring the total gas production, namely measuring the gas production in the fermentation bottle by using a gas pressure converter.
Choline concentration was measured by taking 1.0mL of the fermentation broth, centrifuging at 13000 Xg at 4℃for 2min, and taking the supernatant. Concentration was measured using a bovine choline ELISA assay kit (Emblica Corp., china) and the test procedure was described in the specification.
1.3 Statistical data processing
Data were analyzed using One-way ANOVA (Tukey) in SPSS20.0 software, P <0.05 indicated significant differences.
2. Results
As shown in tables 6 and 7, in the rumen in vitro simulated fermentation, angelic acid (7.2 mmol/L) and 4-cyclopentene-1, 3-dione (1.8-7.2 mmol/L) can inhibit the degradation of choline. Except that angelic acid reduced gas production and 4-cyclopentene-1, 3-dione increased gas production.
TABLE 6 Effect of angelic acid on degradation of choline by rumen in vitro fermentation
Note that the same row of data shoulder marks have no letters indicating that the difference is not significant (P > 0.05), and different letters indicating that the difference is significant (P < 0.05).
TABLE 7 influence of 4-cyclopentene-1, 3-dione on degradation of choline by rumen in vitro fermentation
Note that the same row of data shoulder marks have no letters indicating that the difference is not significant (P > 0.05), and different letters indicating that the difference is significant (P < 0.05).
The results show that the angelic acid and the 4-cyclopentene-1, 3-dione can protect choline in the ration from being degraded by rumen flora, and meanwhile, the 4-cyclopentene-1, 3-dione can also improve the utilization rate of the rumen flora on the substrate.
Claims (4)
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| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN102326714A (en) * | 2011-09-28 | 2012-01-25 | 山东农业大学 | Ruminant rumen by-pass coating material and application thereof in choline coating |
| CN102630817A (en) * | 2011-02-14 | 2012-08-15 | 河北农业大学 | Rumen-protected choline chloride microcapsule additive and preparation method thereof |
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| WO2007037484A1 (en) * | 2005-09-30 | 2007-04-05 | Kaneka Corporation | Curable composition |
| US20080160116A1 (en) * | 2006-12-07 | 2008-07-03 | Herbalscience Singapore Pte. Ltd | Compositions and Methods Comprising Zingiber Species |
| CN102885207B (en) * | 2012-11-07 | 2013-11-06 | 成都大帝汉克生物科技有限公司 | Composite cheese liver flavor attractant and preparation method thereof |
| KR102119178B1 (en) * | 2017-10-12 | 2020-06-04 | 주식회사 이앤사이언스 | Immune-enhancing composition comprising fermented-Angelica gigas extract as effective component |
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| Publication number | Priority date | Publication date | Assignee | Title |
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| CN102630817A (en) * | 2011-02-14 | 2012-08-15 | 河北农业大学 | Rumen-protected choline chloride microcapsule additive and preparation method thereof |
| CN102326714A (en) * | 2011-09-28 | 2012-01-25 | 山东农业大学 | Ruminant rumen by-pass coating material and application thereof in choline coating |
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