CN106265609A - Eugenol application in preparation treatment Salmonella infection medicine - Google Patents
Eugenol application in preparation treatment Salmonella infection medicine Download PDFInfo
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- CN106265609A CN106265609A CN201610935684.3A CN201610935684A CN106265609A CN 106265609 A CN106265609 A CN 106265609A CN 201610935684 A CN201610935684 A CN 201610935684A CN 106265609 A CN106265609 A CN 106265609A
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- eugenol
- salmonella
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- salmonella infection
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- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K31/00—Medicinal preparations containing organic active ingredients
- A61K31/075—Ethers or acetals
- A61K31/085—Ethers or acetals having an ether linkage to aromatic ring nuclear carbon
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- Y—GENERAL TAGGING OF NEW TECHNOLOGICAL DEVELOPMENTS; GENERAL TAGGING OF CROSS-SECTIONAL TECHNOLOGIES SPANNING OVER SEVERAL SECTIONS OF THE IPC; TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
- Y02—TECHNOLOGIES OR APPLICATIONS FOR MITIGATION OR ADAPTATION AGAINST CLIMATE CHANGE
- Y02A—TECHNOLOGIES FOR ADAPTATION TO CLIMATE CHANGE
- Y02A50/00—TECHNOLOGIES FOR ADAPTATION TO CLIMATE CHANGE in human health protection, e.g. against extreme weather
- Y02A50/30—Against vector-borne diseases, e.g. mosquito-borne, fly-borne, tick-borne or waterborne diseases whose impact is exacerbated by climate change
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Abstract
The invention provides eugenol application in preparation treatment Salmonella infection medicine; with human cervical carcinoma cell (Hela) as infection carrier; by celebrating big protective effect, external biochemical test and mice Salmonella infection model all confirm that eugenol, by affecting the function of III type excretory system, thus has significant inhibitory action to Salmonella infection.Antibiotic is during treatment Salmonella infection, the easily generation of in-ductive drug-tolerance, contrast antibiotic therapy, Chinese medicinal compound eugenol can directly suppress the function of Salmonella III type excretory system to play anti-infective effect, and it is difficult to the generation of in-ductive drug-tolerance, has the advantages that cure rate is high.
Description
Technical field
The present invention relates to eugenol application in preparation treatment Salmonella infection medicine, belong to field of medicaments.
Background technology
Eugenol, molecular formula is C10H12O2, it is colourless or pale yellow liquid, has strong Flos Caryophylli fragrance, water insoluble.My god
Right eugenol mainly with the dry flower of the plant clove of myrtaceae rich in Oleum Caryophylli through distill gained volatile oil. warp again
Cross extraction, acidifying, neutralize, the step such as rectification and obtain.Synthetic method prepares eugenol, is raw material with guaiacol, uses pi-allyl
The directly allylation such as chlorine, 1-propenol-3 and obtain.It is mainly used in antibacterial, blood pressure lowering；Can also be used for perfume fragrance and various cosmetic
In product essence and fragrance for detergents formula, it is also possible to for the allotment of edible essence.But have no both at home and abroad at present eugenol for
The report that anti-salmonella infects.
Gram negative bacteria Salmonella comprises 2500 kinds of serotypes, such as Salmonella typhimuriums, Salmonella enteritidis
With salmonella typhi etc..This facultative intracellular bacterial parasite can infect people, animal and plant, has brought huge for the whole world
Big economic loss.The food origin disease occurred most frequently as the mankind, salmonellosis is typically by the water source polluted and food
Caused.The mechanism of Salmonella illustrates an always hot issue.The poison at the pathogenic place of Salmonella stands gene master
On chromosome to be positioned at and plasmid, what those were positioned on chromosome is referred to as pathogenicity island (SPIs) with virulence associated gene.On SPIs
Gene typically there is relatively low G+C content, this is also the mark of SPIs gene, and they for Salmonella evolution have
Play an important role.At present, it has been found that SPIs be more than 10, and SPI-1 is that the ratio wherein studied is more comprehensive.SPI-1
Being positioned at the base of chromosome center 63, by about 40 genomic constitutions, its G+C content is 44.97%.Owing to SPI-1 is in Salmonella
Playing an important role in phagocytic process, it plays the part of very important role in the virulence of Salmonella.The most domestic for sramana
The infection of Salmonella, commonly used antibiotic is treated.But application antibacterials can not shorten the course of disease of patient, promotes on the contrary
Intestinal produces Resistant strain, makes discharge of bacteria time lengthening, causes the difficulty in treatment.The abuse of conventional antibiotic and its can not keep away
The factors such as the easy in-ductive drug-tolerance exempted from result in Salmonella drug resistance in recent years and gradually strengthen, and Antibiotic Resistance is gradually widened.This
The treatment occurring to infect for anti-salmonella clinically of phenomenon makes the matter worse.The most urgently research and develop new mechanism
Anti-infectives prevents and treats Salmonella infection.
Summary of the invention
The molecular structure of eugenol is as follows:
The protectiveness that the present invention is damaged by human cervical carcinoma cell (Hela) is tested and mice Salmonella infection model
Confirm its effect in anti-salmonella infects.
Accompanying drawing explanation
Accompanying drawing 1: the eugenol impact (positive control) on Salmonella three type excretory system function
Accompanying drawing 2: the eugenol impact (negative control) on Salmonella three type excretory system function
Accompanying drawing 3: the eugenol impact (drug treating group) on Salmonella three type excretory system function
Accompanying drawing 4: the eugenol impact analysis to Salmonella growth
Accompanying drawing 5: the eugenol impact on Salmonella internalization
Accompanying drawing 6: eugenol is to Salmonella three type excretory system main effects Protein S ipA quantitative analysis
Accompanying drawing 7: the eugenol protective rate to Salmonella infection mice
Accompanying drawing 8: the weight loss that eugenol suppression Salmonella infection mice causes
Accompanying drawing 9: Salmonella infection mice target organ bacterium colony colonization amount is counted by eugenol
Accompanying drawing 10: the eugenol impact on Salmonella infection mice target organ IL-1 β secretory volume
Accompanying drawing 11: the eugenol impact on Salmonella infection mice target organ IL-6 secretory volume
Accompanying drawing 12: the eugenol impact on Salmonella infection mice target organ TNF-α secretion amount
Detailed description of the invention
1, the detection test of Salmonella three type excretory system function
This experiment is by external structure SL1344-TEM-SipA recombinant bacterium, by eugenol and SL1344-TEM-SipA bacterial strain
Jointly hatch 4h at 37 DEG C of 200rpm, with Hela cell as carrier, add drug treating group and untreated with MOI=50 to cell
In bacterium solution, negative control add DMEM, under the conditions of 37 DEG C infect 30min.Use PBS cell three times, be subsequently added
CCF4 fluorogenic substrate is changed by the color of fluorescence microscope cell, and detection eugenol is to Salmonella three type excretory system
The detection of function.Test result indicate that, eugenol significantly inhibits the function of Salmonella three type excretory system.Eugenol processes
After, the secretory volume of Salmonella three type excretory system effect protein substantially reduces (see Fig. 1, Fig. 2, Fig. 3).
2, the mensuration of growth curve
By Salmonella incubated overnight (37 DEG C, 200rpm), 1:100 amplification culture to OD in LB fluid medium600nm
Value is about 0.1, sets positive group, negative control group and dosing group respectively.Continue to cultivate, and it is each to measure it every 30min
The optical density value of individual sample, until plateau.Test result indicate that: eugenol is dense in the useful effect of suppression Salmonella function
The growth (see figure 4) of Salmonella is had no effect under degree.
3, the test of big protectiveness is celebrated
The final concentration of eugenol of 0,0.1,0.2mM is added respectively in Salmonella SL1344 bacterium solution, and with
SL1344InvA knocks out strain as comparison, 37 DEG C, and 200rpm hatches 4h altogether jointly, and with Hela cell as carrier, MOI=50 infects
Cell, infects 45min under the conditions of 37 DEG C.Use PBS cell three times, in cell culture system, add gentamycin common
Effect 1h, uses PBS cell three times, collects cell and uses Saponin cell lysis, and doubling dilution is applied to solid medium, enters
Row colony counting.Test result indicate that: wild type salmonella invading mechanism under different pharmaceutical concentration processes is gradually reduced, and
Present drug dose dependency (see accompanying drawing 5).
4, three type excretory system effect protein SipA protein immunoblot test
Final concentration of 0,0.1 is added respectively in Salmonella SL1344 bacterium solution, the eugenol of 0.2,0.4mM, 37 DEG C,
200rpm hatches 4h jointly, uses ultraviolet spectrophotometer to measure its OD600Light absorption value, collect identical bacterium amount.Precipitation uses
The SDS-PAGE Buffer of same volume overhangs, and boils 5min subsequently.Deposit sample is used for Western-Blot analyze.Real
Test result to show: eugenol processes and causes SipA protein level to be remarkably decreased.Left figure is the change of endobacillary SipA amount, right figure
For point sample comparison Isocitrate dehydrogenase (ICDH) (see accompanying drawing 6).
5, the therapeutics research of mice Salmonella infection
5.1 set up mice Salmonella infection model
Use streptomycin to process 6-8 week old mice, before infecting, it is prohibited water 6h, use the mode of oral infection to gavage sramana
Salmonella, sets up Salmonella infection animal model.Protective rate test, pathological examination are orally administered to 5 × 107Individual bacterium amount.
5.2 protective rate experiments
With 5 × 107Individual bacterium amount infecting mouse, and 6h gives the eugenol of 50mg/kg body weight and carries out medicine and control after infecting
Treat, average 12h/ time, treat 5 days.After statistics mouse infection Salmonella 24h, 48h, 72h, 96h, 120h, 144h and 168h
Dead number, calculates mortality rate.Result shows, positive group mice died off at 5-8 days respectively, after infection 8 days the most dead
Dying, mortality rate is 100%, and administration group mice respectively the 8th day and the 9th day each dead two, mortality rate is 40%, protective rate
Reach 60%(and seen accompanying drawing 7).
5.3 Avoirdupois monitoring tests
With 5 × 107Individual bacterium amount infecting mouse, and 6h gives the eugenol of 50mg/kg body weight and carries out medicine and control after infecting
Treat, average 12h/ time, treat 5 days.After statistics mouse infection Salmonella 24h, 48h, 72h, 96h, 120h, 144h and 168h
Weight data.Test result indicate that: by analyzing the body weight change of separate groups of mice, after finding to be administered, can significantly alleviate sand
The weight loss (see accompanying drawing 8) of door Salmonella infecting mouse.
5.4 target organ bacterium colony colonization amount counting and enteritis factors checks
With 5 × 107Individual bacterium amount infecting mouse, and 6h gives the eugenol of 50mg/kg body weight and carries out medicine and control after infecting
Treat, average 12h/ time, treat 5 days.Put to death mice in the 6th day, win the caecum of mice, tissue grinder caecal tissue, preparation
The tissue homogenate of 10%, wherein stendomycin flat board counting after partial homogenate liquid doubling dilution, another part homogenate is centrifuged
Remove homogenate supernatant, use ELISA kit to measure IL-1 β, IL-6 and the content of TNF-α in organ lysate.Experimental result
Showing: eugenol drug treating group, in its caecal tissue, the bacterium colony field planting amount of Salmonella substantially reduces (see accompanying drawing 9).Flos Caryophylli
After phenol agents processes, its more positive group of caecal tissue based intracellular cvtokine content significantly reduces (see accompanying drawing 10, Figure 11, Figure 12).
In sum, this research, with Salmonella infection mammalian cell, sets up the medicament protection of cell infection level
Evaluation Model on Effectiveness.Salmonella animal infection modal it has been successfully established in the way of oral infection Salmonella, and it was found that
Eugenol can significantly reduce the mortality rate of Salmonella infection mice, alleviates the enteritis of Salmonella infection mice and suppresses
The decline of infecting mouse body weight, can obviously improve the intestinal inflammation of Salmonella infection mice after showing eugenol treatment.
Claims (3)
1. eugenol application in preparation treatment Salmonella infection medicine.
2. eugenol application in preparation treatment Salmonella infection medicine as claimed in claim 1, it is characterised in that described
Medicine eugenol be pharmaceutically acceptable carrier.
3. eugenol application in preparation treatment Salmonella infection medicine as claimed in claim 1, it is characterised in that described
Salmonella infection refer to by salmonellal human poultry infection, the disease such as including typhoid fever, paratyphoid fever, gastroenteritis and Pullorum Disease
Sick.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201610935684.3A CN106265609A (en) | 2016-11-01 | 2016-11-01 | Eugenol application in preparation treatment Salmonella infection medicine |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
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| CN201610935684.3A CN106265609A (en) | 2016-11-01 | 2016-11-01 | Eugenol application in preparation treatment Salmonella infection medicine |
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| Publication Number | Publication Date |
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| CN106265609A true CN106265609A (en) | 2017-01-04 |
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| Application Number | Title | Priority Date | Filing Date |
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| CN201610935684.3A Pending CN106265609A (en) | 2016-11-01 | 2016-11-01 | Eugenol application in preparation treatment Salmonella infection medicine |
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Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| WO2009151912A1 (en) * | 2008-05-22 | 2009-12-17 | University Of Georgia Research Foundation, Inc. | Antimicrobial composition and use |
| CN104824120A (en) * | 2015-05-11 | 2015-08-12 | 江苏天晟药业有限公司 | Application of eugenol in food-borne pathogenic bacterium in-vitro inhibition |
-
2016
- 2016-11-01 CN CN201610935684.3A patent/CN106265609A/en active Pending
Patent Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| WO2009151912A1 (en) * | 2008-05-22 | 2009-12-17 | University Of Georgia Research Foundation, Inc. | Antimicrobial composition and use |
| CN104824120A (en) * | 2015-05-11 | 2015-08-12 | 江苏天晟药业有限公司 | Application of eugenol in food-borne pathogenic bacterium in-vitro inhibition |
Non-Patent Citations (2)
| Title |
|---|
| INDU UPADHYAYA等: "Efficacy of fumigation with Trans-cinnamaldehyde and eugenol in reducing Salmonella enterica serovar Enteritidis on embryonated egg shells", 《POULTRY SCIENCE》 * |
| 吕世明等: "丁香酚体外抑菌作用研究", 《食品科学》 * |
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Application publication date: 20170104 |