CN108853081A - Medical application of the amentoflavone in treatment chicken necrotizing enterocolitis - Google Patents
Medical application of the amentoflavone in treatment chicken necrotizing enterocolitis Download PDFInfo
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- CN108853081A CN108853081A CN201810613331.0A CN201810613331A CN108853081A CN 108853081 A CN108853081 A CN 108853081A CN 201810613331 A CN201810613331 A CN 201810613331A CN 108853081 A CN108853081 A CN 108853081A
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- amentoflavone
- necrotizing enterocolitis
- perfringens
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- A61K31/00—Medicinal preparations containing organic active ingredients
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- A61K31/35—Heterocyclic compounds having oxygen as the only ring hetero atom, e.g. fungichromin having six-membered rings with one oxygen as the only ring hetero atom
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- A—HUMAN NECESSITIES
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- A61P1/00—Drugs for disorders of the alimentary tract or the digestive system
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- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
- A61P31/00—Antiinfectives, i.e. antibiotics, antiseptics, chemotherapeutics
- A61P31/04—Antibacterial agents
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Abstract
The present invention relates to medical application of the amentoflavone in treatment chicken necrotizing enterocolitis, and inhibit test, biofilm to be formed by C.perfringens sliding motion and test chick necrotic enteritis model therapeutic test, Histopathology sections observation and analysis of intestinal microflora verification experimental verification amentoflavone is inhibited to have protective effect to chicken necrotizing enterocolitis caused by C.perfringens infection.Due to the abuse of conventional antibiotic and the continuous reinforcement of bacterial drug resistance, amentoflavone has cure rate high, it has no drug resistance and the characteristics of medicament residue, therefore amentoflavone is used to treat chicken necrotizing enterocolitis can increase selectivity using drug, to developing new drug important in inhibiting.
Description
Technical field
The invention discloses medical application of the amentoflavone in treatment chicken necrotizing enterocolitis, and it is double yellow to be related to amentotaxus
Application of the ketone in preparation treatment chicken necrotizing enterocolitis drug, belongs to pharmaceutical technology field.
Background technique
C.perfringens is a kind of anaerobic gram positive bacillus, is widely present in soil, the water of nature
It is the normal the gut flora of humans and animals in body, food, human and animal excreta, feed stripped and animal intestinal tract, belongs to conditioned pathogen.
It is different according to the exotoxin of its secretion at present, A-E 5 serotypes altogether are classified as, poultry main infection A type and c-type produce gas pod
Film clostridium, especially based on A type.After C.perfringens infects poultry, mainly cause necrotic enteritis, Symptoms are broiler chicken
Listless, loose random, poor appetite or abolish arrange bloody stool, production performance degradation.Lesion is concentrated mainly on small intestines
Position, it is thinning to show as intestinal wall, has focal or sheet of intestinal mucosa necrosis to fall off.According to related Research statistics, global poultry industry
Because economic loss is up to multi-million dollar caused by necrotic enteritis, the development of poultry farming has been seriously endangered.
Colonization of the C.perfringens in enteron aisle mainly passes through its four types pili (Type IV pili, TFP)
It mediates.TFP is a kind of a kind of locomotive organ being present on bacterial cell membrane, and TFP has C.perfringens at it
The ability of the semisolid dielectric surface movement such as natural habitat and soft or hard agar, this special motion mode are referred to as sliding fortune
Dynamic (Gliding motility).Studies show that, including Colonization, sliding motion, life of the C.perfringens in enteron aisle
The bacteriums such as formation of object envelope function all has certain contact with TFP.By inhibiting the function of C.perfringens TFP, energy
Enough Colonization of the interference bacterium in enteron aisle, to achieve the purpose that anti-C.perfringens infection.
It is the main means for preventing and treating poultry infection and necrosis enteritis by adding antibiotic in feed, however,
Since largely the bacterial drug resistance problem caused by and antibiotic residue problem cause serious prestige to public health to antibiotic
Therefore the side of body develops the hot spot that safely and effectively antibiotic substitute is increasingly becoming research.Native compound have toxicity compared with
The advantages that low, abundance, wide safe range is the valuable source for screening antibiotic substitute.
Amentoflavone is a kind of naturally occurring bis-flavonoid, is primarily present in Amentotaxus plant Honoka
In China fir and Rock lily plant Selaginella tamariscina.Selaginella tamariscina it is civil be referred to as " Herba Selaginellae, it is alive for evermore grass, it is evergreen " etc., tradition doctor
Think that Selaginella tamariscina has the effects of scattered stasis of blood of blood-breaking, blood circulation.Modern medicine study shows that amentoflavone tool is anti-inflammatory, anti-
The biological activities such as cancer, antiviral, antimycotic, anti-oxidant, hypoglycemic, reducing blood lipid.In recent years, a large amount of document is double to amentotaxus
The pharmacological activity of flavones is reported, still, there is no amentoflavone for treating perfringens both at home and abroad so far
The correlative study of chicken necrotizing enterocolitis caused by clostridium.
Summary of the invention
The present invention provides medical application of the amentoflavone in treatment chicken necrotizing enterocolitis, to C.perfringens
Caused chicken necrotizing enterocolitis has good therapeutic effect.
The molecular structure of amentoflavone of the present invention is as shown in Figure 1, molecular formula is：C30H18O10；Molecular weight：
538.46。
The present invention inhibits test, biofilm to be formed and inhibits test, chick necrosis by C.perfringens sliding motion
Property colitis model therapeutic test, Histopathology sections observation and analysis of intestinal microflora verification experimental verification amentoflavone to producing gas
Chicken necrotizing enterocolitis caused by capsular clostridium infection has protective effect：
One, sliding motion inhibits test：
C.perfringens type strain ATCC13124 takes 300 μ l in 37 DEG C of Anaerobic culturel 12h of liquid B HI culture medium later
The culture continues to be incubated in liquid B HI, 37 DEG C of Anaerobic culturel 5h.1ml culture is taken, supernatant, thallus are abandoned in 10000g centrifugation
It is resuspended in 500 μ l BHI fluid nutrient mediums, is allowed to two times of concentrations.The bacterium solution for taking 5 μ l to be concentrated later is added dropwise respectively in preparatory system
It is standby and in 37 DEG C of oven drying 1h containing 4 μ g/ml, the 0.7%BHIA of 16 μ g/ml amentoflavones plate on, 37 DEG C are detested
Oxygen culture 72h is acquired image (referring to fig. 2).The result shows that 4 μ g/ml and 16 μ g/ml amentoflavones can significantly inhibit
Sliding motion of the C.perfringens on BHIA, and 16 μ g/ml amentoflavone function and effect become apparent.
Two, biofilm, which is formed, inhibits test
Supernatant is abandoned in C.perfringens 37 DEG C of Anaerobic culturel 12h in liquid B HI, centrifugation, washed once with sterilizing PBS,
It is resuspended and is adjusted to OD with TSB fluid nutrient medium later600nm=0.1,400 μ l bacterial cultures adjusted are taken, 24 holes are added to
In plate, later with amentoflavone that various concentration is separately added into bacterial cultures so that amentotaxus is double yellow in 24 orifice plates
The final concentration of ketone is respectively 2 μ g/ml, 4 μ g/ml, 8 μ g/ml and 16 μ g/ml, separately sets up addition DMSO and (it is double yellow to be added without amentotaxus
Ketone) positive controls and 400 μ l liquid TSB culture mediums negative control group, 24 orifice plates are placed in 30 DEG C of Anaerobic culturel 5d.It
It inhales afterwards and abandons supernatant, sterilizing PBS is washed twice, and 1% crystal violet, the 400 μ l of filtration sterilization is added in drying at room temperature 30min, incubation at room temperature
30min abandons crystal violet, is washed twice with sterilizing PBS again.33% acetic acid crystal violet is added into every hole, after piping and druming mixes
300 μ l are taken to be separately added into 3 separate wells of 96 orifice plates, every 100 μ l of hole measures light absorption value in microplate reader 570nm, according to crystallization
Purple light absorption value calculates biofilm formation rate.The result shows that amentoflavone can effectively inhibit C.perfringens biological
The formation of envelope (referring to Fig. 3).
Three, the acology research of chick necrotic enteritis model test
1, chick necrotic enteritis model
The male love of 17 ages in days is taken to pull out increasingly (Arbor Acre) Broiler chicks orally administration 0.5ml (1 × 109CFUs it) is resuspended
In the C.perfringens ATCC13124 culture of sterilizing PBS, next day (18 age in days) gives the bacterium solution of identical bacterium amount again to attack
Bacterium establishes chick necrotic enteritis model, and health group broiler chicken orally administration 0.5ml sterilizing PBS is as control.
2, enteron aisle lesion score is tested
Chick orally attacks bacterium and infects preceding 4h (17 age in days) by the amentoflavone of broiler weight orally administration 25mg/kg
(being dissolved in sterilizing PBS), every 12h is administered once.It separately sets up the not positive controls of drug treatment and does not attack the health of bacterium infection
Control group, positive controls weighing same as healthy control group broiler chicken and the sterilizing PBS for pressing weight orally administration respective volume.On
Three groups every group 10 chickens are stated, treatment group is total to drug treatment 5 days (treatment to 21 ages in days).21 age in days cervical dislocations put to death whole meat
Chicken, dissect take duodenum to jejunum, visually observe intestinal lesion, carry out intestinal tract injury according to intestinal tract injury degree and comment
Divide (referring to fig. 4).Intestinal tract injury scoring is using 0-4 points of systems：The tissue of 0 point of expression normal health, without obvious naked eyes lesions visible；
1 point of expression intestinal wall is thinning or frangible, appearance are in canescence；2 points of expression intestinal walls are thinning or frangible, appearance are in greyish white
Color, with focal necrosis, filled with a small amount of gas in intestinal tube；3 points of expression intestinal walls are thinning or frangible, there is a large amount of necrosis region
Or mottled bleeding, enteron aisle are obviously inflated；The serious popularity necrosis of 4 points of expression enteron aisles, with apparent bleeding, fills in enteric cavity
Denounce bulk gas.Above-mentioned small intestine is separately acquired into pathological material of disease, HE dyeing production pathological section, microscopic observation Histopathologic changes (ginseng
See Fig. 5).
When 20 age in days, each group broiler chicken excrement sample is acquired, macro gene order-checking is carried out, it is rich that intestinal flora is analyzed in guiding principle level
Degree variation (referring to Fig. 6).
Enteron aisle lesion score the result shows that, after amentoflavone is treated, significantly reduce necrotic enteritis broiler chicken intestines
The pathological change degree in road.Pathologic examination the result shows that, health group small intestine of broiler chickens intestinal villus structural integrity, no bleeding and
Inflammatory exudation；Control group small intestine of broiler chickens villus necrolysis, structure is imperfect, with bleeding profusely；It is controlled through amentoflavone
After treatment, above-mentioned micropathological damage is relieved.Intestinal flora mutation analysis the result shows that, treated through amentoflavone
Afterwards, clostruidium abundance is substantially less than control group (result only show abundance first three) in enteron aisle, prompts the shuttle in guiding principle level
Shape bacillus Colonization quantity is remarkably decreased.
Based on the above results, it is good to illustrate that amentoflavone has chicken necrotizing enterocolitis caused by C.perfringens
Protective effect.
The positive effect of the present invention is：A kind of new medical application of amentoflavone is provided, it is bad for treating chicken
Dead property enteritis, can significantly reduce the pathological change degree of necrotic enteritis Intestine of Broiler, bad to chicken caused by C.perfringens
Dead property enteritis has good therapeutic effect.
Detailed description of the invention
Fig. 1 is the molecular structure of amentoflavone；
Fig. 2 is that amentoflavone can significantly inhibit sliding motion picture of the C.perfringens on BHIA；
Fig. 3 is the formation figure that amentoflavone can effectively inhibit C.perfringens biofilm；
Fig. 4 is that intestinal tract injury scoring figure is carried out according to intestinal tract injury degree；
Fig. 5 is microscopic observation Histopathologic changes photo；
Fig. 6 is after amentoflavone is treated to the influence diagram for suffering from the horizontal flora abundance of necrotic enteritis Intestine of Broiler guiding principle.
Specific embodiment
Embodiment 1
Application of the amentoflavone in preparation treatment chicken necrotizing enterocolitis drug, is activity system with amentoflavone
At pharmaceutically acceptable any dosage form.
Embodiment 2
Treatment chicken necrotizing enterocolitis described in amentoflavone refers to the necrotic enteritis as caused by bacterium.
Embodiment 3
Treatment chicken necrotizing enterocolitis described in amentoflavone refers to the necrotic enteritis as caused by C.perfringens.
Claims (4)
1. medical application of the amentoflavone in treatment chicken necrotizing enterocolitis.
" medical application of the amentoflavone in treatment chicken necrotizing enterocolitis " 2. as described in claim 1, feature exists
It is that pharmaceutically acceptable any dosage form is made by active constituent of amentoflavone in the amentoflavone.
" medical application of the amentoflavone in treatment chicken necrotizing enterocolitis " 3. as described in claim 1, feature exists
Refer to the necrotic enteritis as caused by bacterium in the chicken necrotizing enterocolitis.
" medical application of the amentoflavone in treatment chicken necrotizing enterocolitis " 4. as claimed in claim 1 or 3, feature
It is that the necrotic enteritis as caused by bacterium refers to the necrotic enteritis as caused by C.perfringens.
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| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201810613331.0A CN108853081B (en) | 2018-06-14 | 2018-06-14 | Application of amentoflavone in preparation of medicine for treating necrotic enteritis of chicken |
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| CN201810613331.0A CN108853081B (en) | 2018-06-14 | 2018-06-14 | Application of amentoflavone in preparation of medicine for treating necrotic enteritis of chicken |
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| CN108853081A true CN108853081A (en) | 2018-11-23 |
| CN108853081B CN108853081B (en) | 2021-04-06 |
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Cited By (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN113143980A (en) * | 2021-01-23 | 2021-07-23 | 吉林大学 | Veterinary selaginella suspension type granule prescription and preparation process |
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2018
- 2018-06-14 CN CN201810613331.0A patent/CN108853081B/en active Active
Cited By (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN113143980A (en) * | 2021-01-23 | 2021-07-23 | 吉林大学 | Veterinary selaginella suspension type granule prescription and preparation process |
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| CN108853081B (en) | 2021-04-06 |
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