CN112741824A - Bacteriostatic agent for inhibiting saprolegnia gymnospirillum in Tibetan plateau - Google Patents
Bacteriostatic agent for inhibiting saprolegnia gymnospirillum in Tibetan plateau Download PDFInfo
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Abstract
The invention provides a bacteriostatic agent for inhibiting water mould of Zygosarijiri in Lasa on the Tibet plateau, which comprises 1-10% of olive extract by mass ratio. The bacteriostatic agent for inhibiting the saprolegnia sicerans of the Tibetan plateau rassa is prepared by combining hydroxytyrosol and oleuropein in an olive extract, has a remarkable bacteriostatic effect, particularly has an excellent bacteriostatic effect on saprolegnia sicerans, and can be used for treating saprolegnia sicerans of the Tibetan plateau rassa.
Description
Technical Field
The invention belongs to the field of aquatic products, and particularly relates to a bacteriostatic agent for inhibiting Aquifex nojirimus in Lasa on the Tibet plateau.
Background
The nude nojirimus (Schizopygopsis), commonly known as native fish, is a scaleless fish, is one of important native economic fishes in the Tibet plateau, belongs to the class of cold water fishes, is mainly distributed in the middle and upper reaches of Yalu Tibetan Yajiangjiang and tributaries and affiliated water bodies thereof, and is a special species in China. The water temperature for culturing the Tibetan plateau Lasa Sejirimus fish is generally within 20 ℃, and the water mold is easy to burst at a lower temperature. And parasites are easily exploded in the fish culture process, and the fish body is damaged due to the breeding of the parasites, so that saprolegniasis is exploded.
The olive leaves are dry leaves of the olives, are large in quantity and belong to wastes and byproducts, past researches show that the olive leaves contain oleuropein, flavone, polyphenol and other bioactive substances, wherein Hydroxytyrosol (HT) can be obtained by converting oleuropein, is a simple phenolic compound in the olives, and has the effects of resisting bacteria, diminishing inflammation, expanding blood vessels, preventing atherosclerosis, resisting cancers, resisting oxidation, inhibiting platelet aggregation and the like.
Oleuropein is a nontoxic phenol secoiridoid compound which is easily absorbed by human body, has the ability of eliminating superoxide anion free radical, hydrogen peroxide, NO and peroxynitroso anion, and also has anticancer, antithrombotic, arteriosclerosis preventing and neuroprotective effects.
Disclosure of Invention
In view of the above, the invention aims to overcome the defects in the prior art and provides a bacteriostatic agent for inhibiting saprolegnia luteorum in the Tibetan plateau lasza.
In order to achieve the purpose, the technical scheme of the invention is realized as follows:
a bacteriostatic agent for inhibiting Aquifex nojirimus of Lasa in Tibet plateau comprises 1-10 wt% of Olea europaea extract.
Preferably, the bacteriostatic agent comprises 3-8% of olive extract by mass.
Further, the olive extract is prepared from the following components in a mass ratio of 1: 0.1-10 parts of hydroxytyrosol and oleuropein.
Preferably, the olive extract is prepared by mixing the following components in a mass ratio of 1: 0.5-8% of hydroxytyrosol and oleuropein.
Preferably, the olive extract is prepared by mixing the following components in a mass ratio of 1: 0.8-5 parts of hydroxytyrosol and oleuropein.
Further, the hydroxytyrosol is prepared by a method comprising the following steps: drying olive leaves, pulverizing into powder, adding the powder into 0.1-0.2mol/L hydrochloric acid solution, heating in water bath for 2-5 hr, adjusting pH value of the solution to neutral, and filtering.
Further, the temperature of the water bath heating is 40-60 ℃; the solid-liquid ratio of the powder to the hydrochloric acid solution is 1: 30-60.
Further, the oleuropein is prepared by the method comprising the following steps: drying Olea europaea leaf, pulverizing into powder, adding into 10-20 times of 60-80% ethanol, extracting at 30-60 deg.C in water bath for 1-3 hr, and filtering.
The application of the bacteriostatic agent, and the application of the bacteriostatic agent in preparing the feed with bacteriostatic effect.
The application of the bacteriostatic agent, and the application of the bacteriostatic agent in preparing a medicament for treating saprolegniasis.
Compared with the prior art, the invention has the following advantages:
the bacteriostatic agent for inhibiting the saprolegnia sicerans of the Tibetan plateau rassa is prepared by combining hydroxytyrosol and oleuropein in an olive extract, has a remarkable bacteriostatic effect, particularly has an excellent bacteriostatic effect on saprolegnia sicerans, and can be used for treating saprolegnia sicerans of the Tibetan plateau rassa.
Detailed Description
Unless defined otherwise, technical terms used in the following examples have the same meanings as commonly understood by one of ordinary skill in the art to which the present invention belongs. The test reagents used in the following examples, unless otherwise specified, are all conventional biochemical reagents; the experimental methods are conventional methods unless otherwise specified.
The present invention will be described in detail with reference to examples.
Example 1 preparation of olive extract
Weighing 100g of olive leaves, drying, crushing into powder, adding the powder into 4L of hydrochloric acid solution with the concentration of 0.12mol/L, heating in water bath at 50 ℃ for 2 hours, adjusting the pH value of the solution to be neutral, and filtering to obtain hydroxytyrosol.
Weighing 100g of olive leaves, drying, pulverizing into powder, adding the powder into 20 times of 80% ethanol, heating and extracting for 2 hours in water bath at 45 deg.C, and filtering to obtain oleuropein.
Weighing 10g of hydroxytyrosol and 10g of oleuropein, and uniformly mixing to obtain the olive extract A.
Weighing 10g of hydroxytyrosol and 40g of oleuropein, and uniformly mixing to obtain an olive extract B.
0.5g of hydroxytyrosol and 10g of oleuropein are weighed and mixed uniformly to obtain the olive extract C.
Weighing 10g hydroxytyrosol and 0.5g oleuropein, and mixing well to obtain an olive extract D.
10g of hydroxytyrosol were weighed to obtain olive extract E.
10g of oleuropein was weighed out to obtain an olive extract F.
EXAMPLE 2 preparation of bacteriostatic Agents
5g of olive extract A is weighed and mixed with 100g of conventional bacteriostatic agent to obtain bacteriostatic agent A.
5g of olive extract B is weighed and mixed with 100g of conventional bacteriostatic agent to obtain bacteriostatic agent B.
5g of olive extract C is weighed and mixed with 100g of conventional bacteriostatic agent to obtain bacteriostatic agent C.
5g of olive extract D was weighed and mixed with 100g of a conventional bacteriostatic agent to obtain bacteriostatic agent D.
5g of olive extract E was weighed and mixed with 100g of conventional bacteriostatic to obtain bacteriostatic E.
5g of olive extract F is weighed and mixed with 100g of a conventional bacteriostatic agent to obtain bacteriostatic agent F.
0.5G of olive extract A was weighed and mixed with 100G of conventional bacteriostatic to obtain bacteriostatic G.
15g of olive extract A was weighed and mixed with 100g of conventional bacteriostatic agent to obtain bacteriostatic agent H.
EXAMPLE 3 bacteriostatic Effect of bacteriostatic agents
And (3) test bacterium culture: activating the strain of saprolegnia, inoculating to solid culture medium, culturing at 15 deg.C for 24 hr, picking out single colony, inoculating to nutrient broth culture medium, and shake culturing for 24 hr. And diluting the bacterial liquid to a proper concentration by adopting a plate counting method, wherein the concentration of the bacterial liquid is 1 multiplied by 101 CFU/mL.
Adding bacteriostatic agent A, bacteriostatic agent B, bacteriostatic agent C, bacteriostatic agent D, bacteriostatic agent E, bacteriostatic agent F, bacteriostatic agent G, bacteriostatic agent H and conventional bacteriostatic agent into different test tubes, and repeating each for 3 times without adding composition as blank control and bacteria as negative control. Shaking (180r/min) in a shaking table at 15 ℃ for 24h, wherein the diameter of the inhibition zone and the inhibition grade are shown in Table 1. The result judgment standard is according to the standard introduced in the Chinese medicine pharmacology: the inhibition zone is more than or equal to 20mm, and the drug is extremely sensitive; the inhibition zone is 15-19mm, belonging to high sensitivity; the zone of inhibition is 10-14mm, belonging to the middle allergy; the inhibition zone is less than 10mm, and the low-sensitivity drug is low-sensitivity drug; has no bacteriostatic zone, and is insensitive (no bacteriostatic effect).
TABLE 1 zone diameter and grade of inhibition
In the above tables, "+ + + + +" indicates extreme sensitivity, "+" indicates medium sensitivity or low sensitivity, and "-" indicates no sensitivity (no bacteriostatic effect).
As can be seen from table 1, bacteriostatic agent a-bacteriostatic agent H all had a bacteriostatic effect on saprolegnia, wherein the bacteriostatic levels of bacteriostatic agent a and bacteriostatic agent B on saprolegnia are extremely sensitive, indicating that the mass ratio of contained is 1: 0.1-10 of hydroxytyrosol and oleuropein, wherein the bacteriostatic agent of the olive extract has obvious bacteriostatic effect on saprolegnia, and the independent use of hydroxytyrosol or oleuropein as bacteriostatic components has obvious difference from the bacteriostatic effect.
Example 4 Effect of bacteriostatic Agents on the Monascus nudivirus in the Tibet plateau
Weighing 100g of olive leaves, drying, crushing into powder, adding the powder into 4L of hydrochloric acid solution with the concentration of 0.12mol/L, heating in water bath at 50 ℃ for 2 hours, adjusting the pH value of the solution to be neutral, and filtering to obtain hydroxytyrosol.
Weighing 100g of olive leaves, drying, pulverizing into powder, adding the powder into 20 times of 80% ethanol, heating and extracting for 2 hours in water bath at 45 deg.C, and filtering to obtain oleuropein.
Weighing 10g of hydroxytyrosol and 10g of oleuropein, and uniformly mixing to obtain the olive extract A.
Weighing 10g of hydroxytyrosol and 60g of oleuropein, and uniformly mixing to obtain an olive extract B.
Weighing 10g of hydroxytyrosol and 2g of oleuropein, and uniformly mixing to obtain an olive extract C.
5g of olive extract A is weighed and mixed with 100g of conventional bacteriostatic agent to obtain bacteriostatic agent A.
5g of olive extract A is weighed and mixed with 100g of conventional bacteriostatic agent to obtain bacteriostatic agent B.
5g of olive extract A was weighed and mixed with 100g of conventional bacteriostatic to obtain bacteriostatic C.
5g of olive extract A is weighed and mixed with 100g of conventional bacteriostatic agent to obtain bacteriostatic agent D.
10g of olive extract A was weighed and mixed with 100g of conventional bacteriostatic to obtain bacteriostatic E.
100g of conventional bacteriostatic agent was weighed as a control group.
In 4-5 months in 2020, acquiring 7000 tails of Tibet native fishes from the field, selecting parents for breeding, and using the rest fishes for water mold bacteriostasis effect tests; the test is carried out in a cement pond with the width of 2m, the length of 10m and the depth of 0.5m, the test is carried out for 7D before the test, after the fish is stable (after the test for 7D, mechanical damage is caused by transportation and selection of the fish, more than 95 percent of the fish has saprolegnia), 600 fish is selected as the test fish and divided into 6 groups, bacteriostatic agent A, bacteriostatic agent B, bacteriostatic agent C, bacteriostatic agent D, bacteriostatic agent E and a control group are respectively added, water inlet and outlet are closed during the administration, and the liquid medicine is drained and then injected with new water after the administration. The mortality, the cure rate and the recurrence rate of Saprolegnia after 7 days of administration of the drugs in each experimental group were counted, and the results are shown in Table 2.
TABLE 2 results of bacteriostats on Zygosarijiri saprolegnia lata in Tibet plateau
| Mortality (%) | Saprolegnia cure rate (%) | 7d Saprolegnia recurrence rate (%) | |
| Bacteriostatic agent A | 2.62±0.34 | 95.41±0.27 | 3.49±0.05 |
| Bacteriostatic agent B | 5.41±0.85 | 96.24±0.74 | 8.64±0.10 |
| Bacteriostatic agent C | 6.74±0.58 | 90.74±0.45 | 8.04±0.25 |
| Bacteriostatic agent D | 10.23±0.84 | 82.41±0.08 | 12.56±0.63 |
| Bacteriostatic agent E | 8.08±0.36 | 80.74±0.92 | 10.58±0.47 |
| Control group | 36.25±0.36 | 53.24±0.72 | 26.74±0.34 |
As can be seen from table 2, bacteriostatic agent a has an outstanding inhibitory effect on watery mold of lasza nojirimus in tibetan plateau, and has significant mortality, cure rate and recurrence rate, indicating that the bacteriostatic agent a comprises the following components in a mass ratio of 1: 0.8-5 parts of hydroxytyrosol and oleuropein, and the bacteriostatic agent containing 3-8% of the above olive extract has remarkable bacteriostatic effect on saprolegnia.
The above description is only for the purpose of illustrating the preferred embodiments of the present invention and is not to be construed as limiting the invention, and any modifications, equivalents, improvements and the like that fall within the spirit and principle of the present invention are intended to be included therein.
Claims (8)
1. A bacteriostatic agent for inhibiting Aquifex nojirimus of Lasa on Tibet plateau is characterized in that: the bacteriostatic agent contains 1-10% of olive extract by mass.
2. The bacteriostatic agent for inhibiting the water mold of Tibetan plateau rassa schizonojirimus, as claimed in claim 1, wherein: the olive extract is prepared from the following components in a mass ratio of 1: 0.1-10 parts of hydroxytyrosol and oleuropein.
3. The bacteriostatic agent for inhibiting the water mold of Tibetan plateau rassa schizonojirimus, as claimed in claim 2, wherein: the olive extract is prepared from the following components in a mass ratio of 1: 0.5-8% of hydroxytyrosol and oleuropein.
4. The bacteriostatic agent for inhibiting the water mold of Tibetan plateau rassa schizonojirimus, as claimed in claim 3, wherein: the hydroxytyrosol is prepared by the method comprising the following steps: drying olive leaves, pulverizing into powder, adding the powder into 0.1-0.2mol/L hydrochloric acid solution, heating in water bath for 2-5 hr, adjusting pH value of the solution to neutral, and filtering.
5. The bacteriostatic agent for inhibiting Seikagajirima nudiflora in Tibet plateau according to claim 4, wherein: the temperature of the water bath heating is 40-60 ℃; the solid-liquid ratio of the powder to the hydrochloric acid solution is 1: 30-60.
6. The bacteriostatic agent for inhibiting the water mold of Tibetan plateau rassa schizonojirimus, as claimed in claim 3, wherein: the oleuropein is prepared by the method comprising the following steps: drying Olea europaea leaf, pulverizing into powder, adding into 10-20 times of 60-80% ethanol, extracting at 30-60 deg.C in water bath for 1-3 hr, and filtering.
7. Use of a bacteriostatic agent according to any one of claims 1-6 wherein: the bacteriostatic agent is applied to preparing the feed with bacteriostatic effect.
8. Use of a bacteriostatic agent according to any one of claims 1-6 wherein: the bacteriostatic agent is applied to the preparation of the medicine for treating saprolegniasis.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN202011638499.0A CN112741824A (en) | 2020-12-31 | 2020-12-31 | Bacteriostatic agent for inhibiting saprolegnia gymnospirillum in Tibetan plateau |
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| Application Number | Priority Date | Filing Date | Title |
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| CN202011638499.0A CN112741824A (en) | 2020-12-31 | 2020-12-31 | Bacteriostatic agent for inhibiting saprolegnia gymnospirillum in Tibetan plateau |
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| CN112741824A true CN112741824A (en) | 2021-05-04 |
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| CN202011638499.0A Pending CN112741824A (en) | 2020-12-31 | 2020-12-31 | Bacteriostatic agent for inhibiting saprolegnia gymnospirillum in Tibetan plateau |
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- 2020-12-31 CN CN202011638499.0A patent/CN112741824A/en active Pending
Non-Patent Citations (3)
| Title |
|---|
| 卫强: "《植物茎叶化学成分的提取分离及活性研究》", 31 December 2018, 合肥：安徽大学出版社 * |
| 李春燕等: "油橄榄叶中羟基酪醇的提取及分离纯化", 《食品与发酵工业》 * |
| 杨德松等: "《农药学实验指导》", 30 November 2016, 中国农业大学出版社 * |
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