CN104856978B - A kind of chitosan spraying film and preparation method thereof - Google Patents
A kind of chitosan spraying film and preparation method thereof Download PDFInfo
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Abstract
The invention provides a kind of chitosan spraying film, it is prepared by the component of following weight percent：Chitosan 0.05~0.5%, lactic acid 0.01%~0.15%, surfactant 0.1%~0.2%, cosolvent 0.1%~0.3%, surplus are water.Present invention also offers the method for preparing the spraying film and the purposes of the spraying film.Chitosan spraying film prepared by the present invention is small to the excitant of skin wound, and good film-forming property, bioavilability is high, strong antibacterial, can effectively facilitate wound healing.
Description
Technical field
The present invention relates to a kind of chitosan spraying film and preparation method thereof, belong to drug field.
Background technology
With the development of world industry, wound, have become disease common in daily life, and wound infection is then
It is the major reason for causing death.Counted according to the World Health Organization, the nineteen ninety various people of the lethal number of wound about 5,100,000 in the whole world,
It is expected that the year two thousand twenty can increase to 8,400,000 people.Wound is a kind of very important disease, therefore carries out treatment and the repairing research of wound
It is very necessary.With the development that Winter, Hinman and Maibaeh etc. wound repair " moistening healing " are theoretical, the surface of a wound
Repair and the application of protection materials auxiliary material gradually grows up.
It is clinical that antibacterial therapy is generally carried out to skin wound using antibiotic medicine dressing, but due to the limitation of its structure,
Though traditional dressing has certain moisture absorption and protective effect, small area is typically only used for, the wound of non-joint is met an urgent need
Treatment, so as to play temporary transient hemostasis, the effect of the surface of a wound is protected, and clinical practice is greatly limited.
Chitosan (chitosan) is that one kind that extraction is formed through deacetylated from crab shell, shrimp shell or the film of fungi is more
Glycan, it is a kind of natural polymer chemical material.Chitosan has hemostasis, antibacterial, promotion wound healing, suppresses cicatrization etc.
A variety of effects, have excellent histocompatbility, inanimate object repellency, do not cause allergic reaction, biodegradability, naturally into
Film, it is excellent medical dressing and tissue engineering material.The chitosan of HMW has good film-forming property, strong antibacterial etc.
Advantage, but because its crystallinity highly dissoluble is poor, can not preparation, using being greatly limited；The chitosan of low molecule amount or
Chitosan derivatives need to prepare by degraded, derivatization method, although its solubility problem is improved, its film forming,
Antibacterial ability has declined.
At present, commercially available and document report chitosan class product be with low-molecular weight chitoglycan (such as
CN102846655A, CN104042719A, CN102824308) or chitosan derivatives (such as CN104586753A) conduct
Active component, although its dissolubility is preferable, antibacterial ability is bad, it is necessary to using heavy dose of active component or adds more
Kind antibacterial substance plays synergy, can be only achieved preferable antibacterial effect.The supplementary material for so resulting in preparation expends
The drawbacks such as excessively, preparation technology complicates, production cost improves, and the quality to preparation and security generation harmful effect.
Meanwhile in order to dissolve chitosan well, chitosan class product uses the relatively good solvent of solubility property at present：Second
Acid, but there is the problem of excitant is strong in it, directly affect the security of medication.
The drawbacks of for existing chitosan class product, preferably supplementary material, control composition content, it is clinical treatment skin trauma
Infection provides a kind of small excitant, good film-forming property, the novel formulation of strong antibacterial, has very important significance.
The content of the invention
The technical scheme is that provide a kind of chitosan spraying film and its production and use.
The invention provides a kind of chitosan spraying film, it is prepared by the component of following weight percentage：Shell
Glycan 0.05-0.5%, lactic acid 0.01%-0.15%, surfactant 0.1%-0.2%, cosolvent 0.1%-0.3%, surplus
For water.
Wherein, described chitosan molecule amount is 10w-30w.
Preferably, described chitosan molecule amount is 30w.
Wherein, described surfactant is n-octyl alcohol, water-soluble silicon oil or dimethicone；
Described solubilizer is Tween 80, benzoic acid or polyethylene glycol 400.
Preferably, the component of percentage by weight is：
Chitosan 0.1%, lactic acid 0.05%, n-octyl alcohol 0.15%, Tween 80 0.25%, surplus are water.
The invention provides the method for preparing chitosan spraying film, comprise the steps：
A. each component is weighed according to proportioning described in claim 1-5 any one；
B. extracting lactic acid, it is prepared into the lactic acid aqueous solution that concentration is 0.1-0.14%；
C. chitosan is taken, is dispersed in the lactic acid solution of b step preparation, is swelled, after stirring and dissolving, regulation pH value to 3.5-
6.0, filtering, obtain chitosan lactic acid solution；
D. cosolvent and surfactant are dissolved in ethanol solution respectively, the ethanol that cosolvent is respectively prepared is molten
The ethanol solution of liquid and surfactant；
E. chitosan lactic acid solution prepared by step c is stirred, adds the ethanol solution of cosolvent made from step c, stirring
To clarification, the ethanol solution of surfactant made from step c is added, is stirred to clarify, it is filling to produce.
Wherein, in described lactic acid aqueous solution, lactic acid concn 0.12%.The ethanol solution and table of described cosolvent
The concentration of the ethanol solution of face activating agent is 5%.
Chitosan of the present invention spraying film to the bacteriostasis rates of Escherichia coli up to more than 99%, the suppression to staphylococcus aureus
Bacterium rate reaches 100%.
Present invention also offers chitosan spraying film to prepare the infection for the treatment of skin trauma and/or wound healing
Medicine in purposes.Described medicine is the medicine of anti-Escherichia coli or staphylococcus aureus.
The present invention is using the poor solvent of solubility property by the particular combination of special component：Under conditions of lactic acid,
The high molecular weight chitosan of dissolubility difference is dissolved well on the contrary, has been made with good film-forming property, strong antibacterial, biological utilisation
High chitosan spraying film is spent, and because without acetic acid is used, its excitant is small, and security is good, and effect experiment proves, this
Invention chitosan spraying film can be effectively antibacterial, treatment skin trauma infection, promotion wound healing, and potential applicability in clinical practice is good
It is good.
Brief description of the drawings
Fig. 1 chitosan spraying film fabricating technology route maps of the present invention
The 0th day traumatic wounds figure of tri- groups of rats of Fig. 2 (is respectively from left to right chitosan spraying film group, commercially available prod A
Group, physiological saline group)
The 7th day traumatic wounds figure of tri- groups of rats of Fig. 3 (is respectively from left to right chitosan spraying film group, commercially available prod A
Group, physiological saline group)
The 14th day traumatic wounds figure of tri- groups of rats of Fig. 4 (is respectively from left to right chitosan spraying film group, commercially available prod A
Group, physiological saline group)
Fig. 5 saline rats histopathology slide (is respectively from left to right No. 2 rat skin section (HE200
×), No. 5 rat skins section (HE200 ×), No. 7 rat skins sections (HE200 ×))
The pathology section of Fig. 6 commercially available prod A groups rat tissue (is respectively from left to right No. 3 rat skin section (HE200
×), No. 6 rat skins section (HE200 ×), No. 8 rat skins sections (HE200 ×))
The pathology section of Fig. 7 chitosans spraying film group rat tissue (is respectively from left to right No. 1 rat skin section
(HE200 ×) No. 4 numbers rat skin section (HE200 ×) No. 9 rat skin section (HE200 ×))
Fig. 8 negative control group rabbit vaginas pathological section (from left to right be respectively rabbit vagina epimere section (HE200 ×),
Rabbit vagina stage casing section (HE200 ×), rabbit vagina hypomere section (HE200 ×))
Fig. 9 chitosans spraying film group control group rabbit vagina pathological section (is respectively from left to right that rabbit vagina epimere is cut
Piece (HE200 ×), rabbit vagina stage casing section (HE200 ×), rabbit vagina hypomere section (HE200 ×))
Below by way of embodiment, the present invention is described in further detail, but is not intended to limit the present invention, ability
The various changes and replacement that field technique personnel make according to the present invention, without departing from the spirit of the present invention, all should belong to this hair
Bright scope of the following claims.
Embodiment
The preparation technology of the chitosan of embodiment 1 spraying film
By 0.05g lactic acid dissolutions in 100g pure water, 0.05% lactic acid solution is made；Weigh 0.2g chitosan (molecules
Measure as 30w), 0.05% lactic acid solution surface swelling 30min is dispersed in, after stirring 1.5h dissolvings, pH to 4.5 is adjusted, filtering, obtains
Chitosan lactic acid solution.Tween 80 0.3g and n-octyl alcohol 0.1g are dissolved in ethanol solution respectively and are made into 5% ethanol solution,
Under stirring, n-octyl alcohol solution is slowly added to micro-sampling pin, adds tween solution, stirred to solution and clarify, it is filling to produce.
Encapsulated immediately after bottling, be that 100,000 grades of workshops are filling in cleanliness factor, sign loading amount 20mL/ bottles should be no less than per bottled amount.(prepare
Processing technology routine figure is shown in Fig. 1)
Application method：Chitosan spraying film is uniformly sprayed apart from surface of a wound 10-15cm position.Sprinkling is pressed every time
Pressure 2-3 times, also can increase or decrease dosage according to patient's actual conditions.Daily to use 2 times, continuous use is one in 15 days and treated
Journey.
The preparation of 2 chitosan of the present invention of embodiment spraying film
By 1g lactic acid dissolutions in 1000g pure water, 0.1% lactic acid solution is made；Weigh 5g chitosan (molecular weight
10W), 0.1% lactic acid solution surface swelling 50min is dispersed in, after stirring 2h dissolvings, pH to 5 is adjusted, filtering, obtains chitosan breast
Acid solution.Tween 80 2g and n-octyl alcohol 1.2g are dissolved in ethanol solution respectively and are made into 5% ethanol solution, under agitation, is used
Micro-sampling pin is slowly added to n-octyl alcohol solution, adds tween solution, stirs to solution and clarifies, filling to produce.
The preparation of 3 chitosan of the present invention of embodiment spraying film
By 5g lactic acid dissolutions in 2500g pure water, 0.2% lactic acid solution is made；Weigh 3.75g chitosan (molecular weight
30W), 0.2% lactic acid solution surface swelling 50min is dispersed in, after stirring 1h dissolvings, pH to 4 is adjusted, filtering, obtains chitosan breast
Acid solution.Tween 80 5g and n-octyl alcohol 4.8g are dissolved in ethanol solution respectively and are made into 5% ethanol solution, under agitation, is used
Micro-sampling pin is slowly added to n-octyl alcohol solution, adds tween solution, stirs to solution and clarifies, filling to produce.
The preparation of 4 chitosan of the present invention of embodiment spraying film
By 10g lactic acid dissolutions in 20kg pure water, 0.05% lactic acid solution is made；Weigh 40g chitosan (molecular weight
30W), 0.05% lactic acid solution surface swelling 1.5h is dispersed in, after stirring 2h dissolvings, pH to 5.3 is adjusted, filtering, obtains chitosan
Lactic acid solution.Tween 80 50g and n-octyl alcohol 36g are dissolved in ethanol solution respectively and are made into 5% ethanol solution, under agitation,
N-octyl alcohol solution is slowly added to micro-sampling pin, adds tween solution, stirs to solution and clarifies, it is filling to produce.
Beneficial effects of the present invention are proved below by way of specific experiment.
Surfactant, the screening test of hydrotropy in 1 chitosan of the present invention of experimental example spraying film
First, the screening of surfactant
1st, experimental method
Chitosan lactic acid solution surface tension value is determined, as the reference index for weighing surfactant.Compound concentration is
0.15% water-soluble silicon oil solution, dimethicone ethanol solution, each 5mL of n-octyl alcohol ethanol solution, above solution is distinguished
The chitosan lactic acid solution for adding 500mL isoconcentrations is configured to three groups of different solution.Under normal temperature and pressure, the solution that will prepare
Circular groove about 1mL among surface tension apparatus measurement ware is implanted sequentially, reading is recorded, repeats every group and be averaged in triplicate
Value.Using surface tension value as evaluation index, optimum surfactant is selected.
2nd, experimental result
The chitosan lactic acid original solution surface tension value of table 1
Note：Experiment is carried out at normal temperatures and pressures, similarly hereinafter.
Influence of the 2 three groups of surfactants of table to the surface tension value of chitosan lactic acid solution
Experiment shows that n-octyl alcohol, water-soluble silicon oil or dimethicone can effectively reduce chitosan lactic acid solution
Surface tension, it may be used to prepare present invention spraying film, wherein, the best results of n-octyl alcohol, it is therefore preferable that n-octyl alcohol is made
For surfactant.
2nd, the screening of cosolvent
1st, experimental method
The polyethylene glycol 400 ethanol solution, Tween 80 ethanol solution, alcohol benzoate solution that compound concentration is 0.25% are each
5mL, above solution is separately added into the chitosan lactic acid solution containing n-octyl alcohol ethanol solution of 500mL isoconcentrations, stirred
10min.Visually observe whether muddiness disappears, and pass through surface tension apparatus at normal temperatures and pressures and determine table after addition cosolvent
The change of face tension force, choose optimal cosolvent.
2nd, experimental result
The pH value situation of change of 3 three groups of chitosan solutions of table
Influence of the 4 three groups of cosolvents of table to the surface tension value of chitosan lactic acid solution
Experiment shows that Tween 80, benzoic acid or polyethylene glycol 400 can effectively reduce the surface of chitosan lactic acid solution
Tension force, it may be used to prepare present invention spraying film, wherein, the best results of Tween 80, helped it is therefore preferable that Tween 80 is used as
Solvent.
3rd, the screening of the proportioning of present invention spraying film
1st, experimental method
Experimental method：In the case of above-mentioned preferable n-octyl alcohol, Tween 80, fixed chitosan dosage, prepare difference and match somebody with somebody
Than Tween 80 and n-octyl alcohol series concentration sample, using surface tension as Testing index, commercial preparation is reference preparation, and screening is optimal
Proportioning, specific experiment result see the table below.
The different auxiliary of table 5 than auxiliary material screening test
Before cosolvent is not added, group 3-8 sample solutions have white flock precipitate, and liquid is muddy, in group 9-12 samples
Various concentrations n-octyl alcohol ethanol solution is added in product, liquid clarity is good, especially with group 12, and group 12 match under it is molten
Liquid surface tension is minimum, therefore the optimum proportioning of present invention spraying film is the tween second of 0.15% n-octyl alcohol ethanol solution+0.25%
The chitosan lactic acid solution of alcoholic solution+0.1%.
2 chitosan of the present invention of experimental example spraying film film-formation result double blind experiment
(1) experiment material：
Chitosan spraying film (being prepared by embodiment 3) of the present invention, commercially available prod A (Jin Fusheng, Guangzhou profit rainbow medical sci-tech
Co., Ltd), commercially available prod B (gold wound chitosan wound care film, Hubei Pu Ai pharmaceutcal corporation, Ltds)
(2) experimental method：
6 subjects participate in double blind experiments, and three kinds of sprays are sprayed on skin, using mobility, drying property, film forming as into
The evaluation index of film effect.Wherein mobility standards of grading are：1 point-be very easy to be lost in, 2 points-be easier to flowing, 3 points-very
Flow less, 4 points-do not flow completely；Drying property standards of grading are：1 point is drying time>7min, 2 points of drying times are 5-
7min, 3 points of drying times are 3-5min, 4 points of drying times<3min；Film forming standards of grading are：1 point-imperceptible obvious
Film, 2 points-can be formed incomplete film, 3 points-can form obvious complete film.Add up all fractions, and calculates all tested
The total score (± SD) of person, score value is more high then to represent that film-formation result is better.
(3) experimental result：
The double blind experiment test result of table 6
The film-formation result appraisal result of table 7
In double blind experiment it can be seen from film-formation result appraisal result the moulding property of chitosan of the present invention spraying film it is good,
Mobility is small, and its film-formation result is better than gold wound chitosan nursing film and Jin Fusheng chitosan antibacterial filming sprayers.
Experimental result illustrates, the moulding property of chitosan of the present invention spraying film is good, mobility is small, is significantly better than existing commercially available
Product, bacterium can be effectively obstructed, promote wound healing, treatment skin trauma infection.
3 chitosan of the present invention of embodiment spraying film film forming surface tension, film formation time detection
(1) experiment material：
Chitosan spraying film (being prepared by embodiment 3) of the present invention, commercially available prod A (Jin Fusheng, Guangzhou profit rainbow medical sci-tech
Co., Ltd)
(2) experimental method：
A. film formation time detection method：It is 20-25 DEG C, under conditions of relative humidity 70% in temperature, about (will presses 0.2mL
Pressure is twice) decoction is sprayed on the glass plate of dried and clean curtain coating at distance 5-10cm and paves, spontaneously dry.
B. surface tension test experimental method：Determine the chitosan spraying film under the conditions of the optimum proportioning described in embodiment 5
Agent and commercially available prod A surface tension values, as the reference index for weighing surfactant.Under normal temperature and pressure, water, chitosan are sprayed
Mist film and commercially available prod solution A are implanted sequentially circular groove about 1mL among surface tension apparatus measurement ware, record reading,
Every group is repeated to average in triplicate.
(3) experimental result：
The film formation time testing result of table 8
The surface tension contrasting detection result of table 9
Chitosan spraying film is small than commercially available prod A surface tension it can be seen from experimental result, film formation time
Short, rate of film build is fast.
Experimental result illustrates that the film formation time of chitosan spraying film of the present invention is short, and rate of film build is fast, is significantly better than existing
Commercially available prod, bacterium can be effectively obstructed, promote wound healing, treatment skin trauma infection.
4. chitosan of the present invention of experimental example spraying film promotes the test of pesticide effectiveness of wound healing
(1) experiment material
Experiment reagent：Chitosan spraying film (being prepared by embodiment 3)；10% chloraldurate solution；Physiological saline；It is commercially available
Product A (Jin Fusheng chitosan antibacterial filming sprayers, Guangzhou Run Hong Pharmaceutical Technology Co., Ltd, 30mL/ bottles)；Medicinal alcohol.
Experimental animal：SD rats 30, (it is purchased from Jiangning county Qinglongshan animal reproduction field, animal productiong licensing
Number：SCXK (Soviet Union) 2010-0005), male and female half and half, 6-8 week old.Buy within 1 week before experiment, put 25-27 DEG C of temperature, the animal of sound insulation
Interior, sub-cage rearing, freely ingest, drink water.
(2) experimental method：
Model is established：30 rats, it is randomly divided into 3 groups, every group 10.Be divided into chitosan spray group, physiological saline group,
Commercially available prod A groups.10% chloraldurate 0.7m is injected intraperitoneally, the hair for shaving off back about 3cm × 3cm sizes is pushed away using electricity,
The region for marking 2cm × 2cm after 75% ethanol disinfection with marking pen is sprayed, back skin of unhairing is cut off with eye scissors, dries 1h
After record wound area, each group gives corresponding laboratory sample respectively, and decoction sprays (pressing is twice) at distance 5-10cm, daily
Administration.
(3) Testing index：The wound situation of observation rat wound site daily, and after surgery 3,5,7,9,11,14,17,
21 days dynamic observation Rat Wound Healing situations, measure surface of a wound area and calculate healing rate, healing rate=(original face area-existing
Surface of a wound area)/original face area, it is considered that traumatic wounds dry tack free is not suppurated, and scab comes off naturally, wound in it is fair and tender just
Normal skin shape is considered as healing.
(4) experimental result
Wound healing situation is shown in Fig. 2,3,4.
Wound area healing state table：
The chitosan of table 10 spraying film group wound area
The physiological saline group wound area of table 11
The commercially available prod A group wound areas of table 12
13 3 groups of Rat Wound Healing rates of table
(note：Compared with chitosan spray group, * is P ＜ 0.05, and * * are P ＜ 0.01.)
From three groups of Rat Wound Healing rates：Compared with physiological saline, chitosan of the present invention sprays film in the prometaphase
Healing Rate all significantly improve, difference extremely significantly (P ＜ 0.01),；Compared with the A of commercially available prod, chitosan spraying film of the present invention
Agent significantly improves in the Healing Rate of mid-term, significant difference (P ＜ 0.05).
Experimental result illustrates, chitosan of the present invention spraying film wound healing, the effect for the treatment of skin trauma infection
It is good, it is significantly better than existing commercially available prod.
5 chitosan of the present invention of experimental example spraying film is tested to the histopathological study of rat
(1) experiment material
Experimental animal：Take each 3 of every group of the rat tested in experimental example 4 the 21st day at random.
Experiment reagent：Chitosan spraying film；10% formalin；Physiological saline；Commercially available prod A (Jin Fusheng chitosans
Antibacterial film forming spray, Guangzhou Run Hong Pharmaceutical Technology Co., Ltd, 30mL/ bottles)；Medicinal alcohol；
(2) experimental method：It is random to put to death 3 rats of each group, separate wound part and its surrounding skin tissue, 10% Fu Er
Malin fixes, and draws materials, dehydration, FFPE.Section is dyed through HE (hematoxylin eosin staining), and observation by light microscope is simultaneously commented
Point.
(3) standards of grading：
A. epidermal cell whether there is degeneration necrosis, forms rotten to the corn, ulcer, crust：It is chosen as 0 point -4 points.0 point without necrosis, by light
It it is 1 point -4 points to weight；(eyepiece 4 ×, object lens 10 ×) measurement under 40 times of light microscopics is burst with " the micro- chi in microscope side " for downright bad length
The length (cm/40 times) of ulcer；
B. corium and hypodermis blood vessel whether there is expansion, hyperemia, oedema, cell infiltration below slough：It is chosen as 0
Divide -4 points.0 point without obvious lesion, 1 point of -4 points of lesion degree is respectively slight, moderate, severe, pole severe；
C. granulation tissue hyperplasia or fibrosis：It is divided into 0-4 points, 0 point：Ripe granulation tissue, complete fibrosis；1 point：Moderate
Maturation, blood vessel inflammatory cell significantly reduces in granulation tissue, the obvious hyperplasia of fibroblast；2 points：It is slight ripe, in granulation tissue
Vasculitis Leukopenia, fibroblast proliferation；3 points：Granulation tissue is in early stage in inmature stage, fibrosis unobvious；4 points：
Without granulation tissue, without fibroblast proliferation；
D. damage surrounding epithelial tissue whether there is hyperplasia：It is divided into 0-3 points.0 point：Damage is local be entirely regeneration, differentiation it is good
Good epithelial tissue covering；1 point：Part has epithelial tissue regeneration covering, the undifferentiated maturation of epithelium；2 points：There is epithelial tissue part
Regeneration；3 points：Without epithelium regeneration.
(4) experimental result
Pathology section examination：See Fig. 5,6,7.
Physiological saline group：In 3 skin histologies, there is ulcer on 2 surfaces, and another 1 original damage location is by complete
Skin group covers, No. 2, No. 7 lesion degrees it is similar, rat part epidermis, corium holostrome missing, form ulcer, length is respectively
There is the inflammatory necrosis tissue of volume on 0.7cm or 1.9m/x40, surface, the granulation tissue of its lower visible hyperplasia, there is the capillary of new life
Blood vessel and fibroblast composition, inside have cell infiltration, inflammatory cell type is mainly mononuclear macrophage and granulocyte.No. 5
Injured surface is covered by intact skin, angling that epidermis is slight, and corium is filled, blood in granulation tissue without annex by granulation tissue
Pipe and inflammatory cell quantity are reduced, fibroblast proliferation.
Commercially available prod A groups：Lesion degree compared with physiological saline group mitigate, No. 6, No. 8 lesion degrees it is close, rat part table
Skin, corium holostrome, which lack, forms ulcer, and length is respectively 1.0cm or 1.8cm/x40, and there is inflammatory necrosis tissue on surface, visible under it
The granulation tissue of hyperplasia, it is made up of the capillary and fibroblast of new life, inside there is a cell infiltration, inflammatory cell type is same
Before.No. 3 injured surfaces are covered by intact skin, angling that epidermis is slight, and corium is filled, granulation group without annex by granulation tissue
Knit interior blood vessel and inflammatory cell quantity to reduce, fibroblast proliferation.
Chitosan spraying film group：The damage of this group is most light, No. 4 rat part epidermises, corium holostrome missing, forms ulcer,
Length:2.9cm/x40, surface are inflammatory necrosis tissue, the granulation tissue of its lower visible hyperplasia, have new life capillary and into
Fibrocyte forms, and inside has a cell infiltration, and inflammatory cell type is the same.No. 1 injured surface is covered by intact skin, and surface has
Slight angling, corium are filled, blood vessel and inflammatory cell quantity are reduced in granulation tissue, fibroblast without annex by granulation tissue
It is more.No. 9 lesion degrees are lighter than No. 1, and granulation ulcer bottom without inflammatory cell, no blood vessel, is filled by the collagenous fibres of maturation substantially.
Lesion score：It is shown in Table 14-17.
The physiological saline group pathological score table of table 14
The commercially available prod A group pathological score tables of table 15
The chitosan of table 16 spraying film group pathological score table
The rat skin lesion light and heavy degree appraisal result of table 17
(5) experiment conclusion
As can be seen that compared with the A groups of commercially available prod, the healing of chitosan spraying film of the present invention is more preferable：Commercially available production
The local epidermis of product A groups, corium holostrome missing, form ulcer, and inflammatory necrosis tissue of the skin surface covered with volume is visible under it
Granulation tissue hyperplasia.Granulation tissue is made up of the capillary and fibroblast of new life, inside there is cell infiltration.Shell of the present invention
Damage topical ulcers have been repaired in glycan spraying film group, and surface covers complete epithelium, and upper subcutaneous granulation tissue is more ripe, fine
Dimensional tissue increases, and blood vessel number and inflammatory cell quantity are reduced.Experimental result illustrates that chitosan spraying film of the present invention can promote
Wound healing, treatment skin trauma infection, and significant effect is better than existing commercially available prod.
6 chitosan of the present invention of experimental example sprays film to rabbit vagina mucous membrane irritation test
(1) experiment material
Experiment reagent：Chitosan spraying film (being prepared by embodiment 3), physiological saline, medicinal alcohol.
Experimental animal：First Adult female unpregnancy New Zealand experimental rabbit 6 (is purchased from Jiangning county's Qinglongshan animal reproduction
, animal productiong licensing number：SCXK (Soviet Union) 2010-0022), body weight 2.6-3.0kg, animal is in estrus, vagina
Mouthful without secretion, without congestion and edema and other damages.Buy, raised in regular grade Animal House, temperature 20-26 in experiment the last week
DEG C, 60 decibels of relative humidity 40-70%, noise ＜, drinking-water of freely ingesting.
(2) experimental method
Model is established：Experimental animal is randomly divided into chitosan spraying film group and saline control group, every group 3.
Syringe is connected with homemade flexible pipe (the mellow and full no corner angle in flexible pipe head), being inhaled in syringe has sample liquid, and flexible pipe is gently inserted
Enter vagina 4-5cm, inject sample liquid, and slowly extract flexible pipe out.Successive administration 10 days.In 24h after processing in 10th day, put to death real
Test animal, cut open the belly take out complete vagina be placed in 10% formalin it is fixed, it is fixed fully after, in vagina epimere, stage casing and
Hypomere is drawn materials respectively, routine paraffin wax embedding, HE (hematoxylin-eosin) dyeing, and om observation vaginal mucosal epithelium cell whether there is change
Property, necrosis, whether there is cell infiltration, it is upper subcutaneous to whether there is congested, oedema and cell infiltration.
Standards of grading：According to the degree of each site morbidity from light to heavy, quantification of slight or very small amount " 0.5 point " successively,
Slightly or on a small quantity " 1 point ", moderate or more " 2 points ", severe or volume " 3 points ", pole severe or a large amount of " 4 points ", normal " 0 point " tired
Add all fractions, and calculate dividing equally for every group of every animalScore value is higher to represent that degree of injury is more serious.
(3) experimental result
Pathology section examination：See Fig. 8, Fig. 9.
Saline control group pathology diagosis：Visible rabbit vagina epimere and stage casing mucous membrane are coated on simple columnar under light microscopic
Skin, lamina propria are the connective tissue rich in blood vessel, and deep is connective tissue and smooth muscle.Hypomere mucous epithelium layer thickens, part
For stratified squamous epithelium.Specific lesion is as follows：
Epimere：3 epimere mucomembranous epithelial cells are shown in very small amount or few without obvious denaturation, necrosis, lamina propria mild hyperaemia
Amount (slight or slight) cell infiltration, inflammatory cell type is mainly neutrophil leucocyte and mononuclear macrophage.It is other to have no bright
Aobvious lesion.Stage casing：3 mucomembranous epithelial cells are slightly denatured, and have a small amount of, very small amount or moderate to be same as above the inflammatory cell leaching of type
Profit；Proper mucous membrane moderate or mild hyperaemia, and see very small amount (slight) cell infiltration, inflammatory cell type is the same.Hypomere：Disease
Change degree overweights epimere and 3, stage casing mucomembranous epithelial cell is slight or moderate is denatured, and has the inflammation that a small amount of or moderate is same as above type
Cellular infiltration；Proper mucous membrane moderate or mild hyperaemia, oedema, and see the cell infiltration of moderate or slight the same type.
Chitosan spraying film group pathology diagosis：Lesion degree is lighter than saline control group, specific as follows：
Epimere：3 epimere proper mucous membranes are slight or slight congested, other to have no obvious lesion.Stage casing：On 3 mucous membranes
Chrotoplast is slightly denatured, wherein the cell infiltration of 1 the same type of degree of taking a favourable turn.It is proper mucous membrane moderate, slight or slightly fill
Blood, 1 with slight cell infiltration.Hypomere：3 mucomembranous epithelial cells are slightly denatured, with the inflammation of slight or slight the same type
Cellular infiltration.3 proper mucous membrane moderates or mild hyperaemia, with slight cell infiltration.
The chitosan spray group rabbit vagina of table 18 section pathological score table
Note 1：Numeral shows lesion degree：0.5 point (slight or very small amount), 1 point (slight or a small amount of), 2 points (moderate is medium
Amount), 3 points (severe or volumes), 4 points (pole severe is a large amount of)
The physiological saline group rabbit vagina of table 19 section pathological score table
The rabbit vaginopathy of table 20 becomes light and heavy degree appraisal result
From experimental result, negative control group vaginal mucosal epithelium cell is slightly denatured, has cell infiltration.Lamina propria
Congested, oedema is with same type of cell infiltration.It is most obvious with hypomere lesion in 3 sections of upper, middle and lower.With chitosan spray
The above-mentioned lesion degree of vagina mucosa is relatively light after administration, illustrates that stimulation of the chitosan spraying film of the present invention to vagina mucosa is made
With very small.Compared to for similar-type products using acetic acid as solvent, the present invention uses endogenous material lactic acid instead on the market at present
As solvent, skin irritation is reduced, in addition it is more preferable than the security of physiological saline.
Experimental result illustrates that the security of chitosan spraying film of the present invention is good, and physiological saline is compared to skin irritation
It is also small.
7 chitosan of the present invention of experimental example spraying film bacteriostatic test
(1) experiment material：
Chitosan spraying film (being prepared by embodiment 3, lot number 3,8,14,18,19,24,29,32) of the present invention；Physiology
Salt solution；Culture medium：Nutrient broth medium (the medicine scientific and technological development company of Beijing three) is prepared by formula, adjusts pH6.5, is added
1.2% agar, after autoclaving, 45 DEG C of constant temperature water baths are standby.Experimental strain：Selection standard strain Escherichia coli ATCC25922,
Staphylococcus aureus ATCC6538, Candida albicans ATCC10231, is stored in -80 DEG C.It is prepared by bacteria suspension：Take each test organisms
The fresh slant strains of strain, lower lawn is washed with sterile saline, with Maxwell opacity tube than turbid, be suitably diluted to concentration about 1-9 ×
106cfumL-1 is standby.
(2) experimental method：
Bacteriostasis rate determines：Take each sample 5mL to be directly added into sterile test tube respectively, prepare 3 parts：Respectively in each test tube plus
Entering a kind of experiment bacteria suspension 0.5mL, make bacterial concentration be about 1-9 × 105cfumL-1, DL instrument mixes, place 0 in 25 DEG C,
2nd, 10, take 0.5mL to add physiological saline after 20min and suitably dilute the counting of rear plate method.It is simultaneously same using physiological saline as control
Method determines.Calculate bacteriostasis rate：Bacteriostasis rate %=(A-B)/A × 100%A：Physiological saline negative control count results B：Sample meter
Number result
(3) experimental result：
The Escherichia coli bacteriostasis rate measurement result of table 21
The staphylococcus aureus bacteriostasis rate measurement result of table 22
The Candida albicans bacteriostasis rate measurement result of table 23
(4) experiment conclusion
According to《Disposable Sanitary Accessory sanitary standard (GB15979-2002)》In " C4：Stripping property antimicrobial product presses down
Evaluation criterion in bacterium method for testing performance ", chitosan spray film to Escherichia coli, staphylococcus aureus and Candida albicans
Stronger bacteriostasis is shown, wherein more than 99% is reached to the inhibiting rate of Escherichia coli, the suppression to staphylococcus aureus
Rate reaches 100%.
Experimental result illustrates that chitosan spraying film bacteriostasis of the present invention is strong, particularly to clinical common pathogenic bacteria：Greatly
Enterobacteria, staphylococcus aureus and Candida albicans have clear and definite inhibitory action, can treat infectious caused by them
Disease, treatment skin trauma infection.
Claims (3)
- The film 1. a kind of chitosan is sprayed, it is characterised in that：Its component is chitosan, lactic acid, n-octyl alcohol, tween and water；Its In per 2500g water in, contain 3.75g chitosans, 5g lactic acid, 4.8g n-octyl alcohols and 5g Tween 80s；Wherein, described chitosan point Son amount is 30w；It is prepared by following methods：A. each component is weighed according to above-mentioned weight；B. extracting lactic acid, it is prepared into the lactic acid aqueous solution that concentration is 0.2%；C. take chitosan, be dispersed in b step preparation lactic acid solution in, be swelled 50min, stirring 1h dissolving after, regulation pH value to 4, filtering, obtain chitosan lactic acid solution；D. Tween 80 and n-octyl alcohol are dissolved in respectively be made into ethanol solution 5% ethanol solution；E. chitosan lactic acid solution prepared by step c is stirred, n-octyl alcohol solution made from addition step d, step d is added and is made Tween 80 solution, stir to clarify, it is filling to produce.
- 2. the spraying film described in claim 1 is in the medicine of the infection for the treatment of skin trauma and/or wound healing is prepared Purposes.
- 3. purposes according to claim 2, it is characterised in that：Described medicine is anti-Escherichia coli or Staphylococcus aureus The medicine of bacterium.
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| CN108498531A (en) * | 2018-06-01 | 2018-09-07 | 广东药科大学 | A kind of chitosan oral solution and preparation method thereof |
| US20210401765A1 (en) * | 2018-10-27 | 2021-12-30 | Shilpa Medicare Ltd | Spray compositions of chitosan for wound healing |
| CN111718687B (en) * | 2019-12-10 | 2023-04-25 | 杭州惠康医疗器械有限公司 | Medical mirror anti-fog liquid and preparation method thereof, medical mirror anti-fog film and preparation method thereof, anti-fog method and medical instrument |
| CN112438964A (en) * | 2020-11-30 | 2021-03-05 | 威兰德(山东)生物科技有限公司 | Composite antibacterial film spraying agent of lysostaphin and chitosan |
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| CN101278948A (en) * | 2008-05-23 | 2008-10-08 | 大连大学 | Biological medical membrane and method of preparing the same |
| CN102670929A (en) * | 2012-06-01 | 2012-09-19 | 山东卫康生物医药科技有限公司 | Composite for injury healing and preparing method thereof |
| CN102824308A (en) * | 2012-08-31 | 2012-12-19 | 广州润虹医药科技有限公司 | Chitosan antibacterial filming sprayer and preparation method |
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| CN101278948A (en) * | 2008-05-23 | 2008-10-08 | 大连大学 | Biological medical membrane and method of preparing the same |
| CN102670929A (en) * | 2012-06-01 | 2012-09-19 | 山东卫康生物医药科技有限公司 | Composite for injury healing and preparing method thereof |
| CN102824308A (en) * | 2012-08-31 | 2012-12-19 | 广州润虹医药科技有限公司 | Chitosan antibacterial filming sprayer and preparation method |
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