CN103893816A - Method for preparing composite amniotic extracellular matrix dressing of bacterial cellulose containing plant ingredients - Google Patents
Method for preparing composite amniotic extracellular matrix dressing of bacterial cellulose containing plant ingredients Download PDFInfo
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Abstract
The invention discloses a method for preparing composite amniotic extracellular matrix dressing of bacterial cellulose containing plant ingredients, and relates to a technology for preparing a medical material. The method comprises the following steps: selecting a bacterial strain capable of secreting bacterial cellulose to activate and prepare seed mash; then mixing the seed mash of which the bacterial strain concentration is 30-50wt% with culture medium containing natural rosemary freeze-dried powder and then carrying out static culture in a culture container to obtain a bacterial cellulose membrane containing natural plant ingredients; taking amniotic extracellular matrix to soak into the culture medium; paving on the surface of the bacterial cellulose membrane after completely wetting; continuing to carry out static culture for 1-6 hours; carrying out purification and sterilization to obtain the composite amniotic extracellular matrix dressing of the bacterial cellulose containing the natural plant ingredients. The method is simple and feasible in preparation process, low in cost, and wide in material source, and the prepared dressing has the properties, such as sterilization, moisturizing, absorbing wound exudates, accelerating wound healing and the like, and is applicable to various chronic skin wounds.
Description
Technical field
The present invention relates to the preparing technical field of medical material, refer to especially the preparation method of the compound characteristics of amniotic extracellular matrix dressing of a kind of Bacterial cellulose that contains natural plant composition.
Background technology
Amniotic membrane is from cytotrophoblast derivation, is the internal layer of two-layer fetal membrane.Normal amniotic membrane is thin and bright, does not contain the structures such as blood vessel, nerve and lymphatic vessel, and thickness is 20～500 μ m.The amniotic membrane tool structure that is of five storeys can be divided into: epithelial layer, basement membrane layer, compacted zone, fibroblast layer, spongy layer.Remove in the characteristics of amniotic extracellular matrix material after cell and contain the Multiple components such as collagen, glycoprotein, proteoglycan, integrin and laminated body, express the associated protein of multiple somatomedin and mRNA thereof, can, for the propagation of cell, differentiation provide abundant nutritional labeling, be conducive to the growth and breeding of cell; Basement membrane layer is thicker, is conducive to attaching and the migration of epidermis cell, promotes epithelization, prevents its apoptosis; The collagen fiber of compacted zone and spongy layer form loose 3 D stereo configuration; Histocompatibility is good, does not express human leukocyte antigen, without induction rejection; Easily obtain, and plasticity is good, is easy to processed.Studies confirm that in a large number, amniotic membrane has good effect as biological dressing for skin injurys such as treatment large-area burns, scalds.But because amniotic membrane is thin and soft, insulation poor water retention property, wound surface ooze out and mostly be that Yi Rong loses, biomechanical property is poor simultaneously, operation acquires a certain degree of difficulty, and it is restricted in biological dressing is applied.
Bacterial cellulose is a kind of natural biopolymer, has hyperfine network structure, is combined into the thick fibre bundle of 40～60 nanometers by the fento of diameter 3～4 nanometers, and is intertwined to form flourishing hyperfine network structure.And its chemical purity is very high, high mechanical strength, good fluid absorbent and performance of keeping humidity while thering is good biocompatibility, hygrometric state due to Bacterial cellulose, make it in medical biotechnology Materials, show great advantage, can be used as desirable degradable carrier material and be applied to the numerous areas such as wound dressing, drug carrier material.The simple Bacterial cellulose of external employing has been reported as moist dressing, and industrialization is for clinical.
Herba Rosmarini Officinalis (Rosmarinus officinatis L.) has another name called You Ancao, and another name tansy, is Labiatae Rosmarinus plant, perennial evergreen undershrub.Most important in Herba Rosmarini Officinalis extract is rosemary antioxidant and rosemary ethereal oil.Rosemary natural anti-oxidant Stability Analysis of Structures, is not easy to decompose, resistance to 190 ℃ of high temperature, and its antioxidant effect is good, and does not have toxicity.In rosemary plant, contain the multiple compounds such as phenol, acid, ketone, terpene, food, medicine, industrial being widely used.Herba Rosmarini Officinalis extract has good anti-microbial property, and staphylococcus aureus, escherichia coli, bacillus subtilis, Hansenula anomala bacterium, penicillium sp, aspergillus niger, Aspergillus flavus and Aspergillus citrimum are all had to good inhibition; Herba Rosmarini Officinalis extract is used for the treatment of chronic wounds and can reduces inflammation, strengthens wound contraction, contribute to the reconstruction of epithelial tissue and granulation tissue, and the formation that can improve wound vascularization and collagen protein.In sum, the Bacterial cellulose that contains natural Herba Rosmarini Officinalis extract composition is combined with characteristics of amniotic extracellular matrix material and obtains having the composite of three-dimensional net structure, can prepare the moist dressing that has histocompatibility, antibacterial, moisturizing, absorbing wound exudate, accelerating wound healing and satisfactory mechanical property concurrently.
This patent adopts the compound characteristics of amniotic extracellular matrix material of microorganism fermentation culture method original position.The bacteria cellulose material that contains natural Herba Rosmarini Officinalis extract composition and characteristics of amniotic extracellular matrix material, collagen fiber interaction and nanofiber by Bacterial cellulose nanofiber and characteristics of amniotic extracellular matrix material compacted zone and spongy layer run through mutually, form and link composite closely.Preparation is simple in the present invention, cost is low, material source is extensive, and the dressing of preparation has the performances such as sterilization, moisturizing, absorbing wound exudate and accelerating wound healing, is applicable to all kinds of chronic skin wounds.
Summary of the invention
The object of this invention is to provide the preparation method of the compound characteristics of amniotic extracellular matrix dressing of a kind of Bacterial cellulose that contains natural plant composition.Relate to a kind of technology of preparing of medical material.Preparation is simple in the present invention, cost is low, material source is extensive, the dressing of preparation has the performances such as sterilization, moisturizing, absorbing wound exudate and accelerating wound healing, is applicable to all kinds of chronic skin wounds such as diabetic foot ulcer, venous leg ulcers, pressure ulcer, skin donor site wound.
The invention discloses the preparation method of the compound characteristics of amniotic extracellular matrix dressing of a kind of Bacterial cellulose that contains natural plant composition, comprise: choose can secreting bacteria cellulosic bacterial strain activation and be prepared into seed mash, the seed mash that is then 30～50wt% by bacterial strain concentration mixes with the fermentation medium that contains natural Herba Rosmarini Officinalis lyophilized powder and is placed on standing cultivation in culture vessel and obtains the bacteria cellulose film that contains natural plant composition; Get characteristics of amniotic extracellular matrix film and be immersed in culture medium, after its complete wetting, be laid in bacteria cellulose film upper surface; Continue to leave standstill and cultivate 1～6h, purified processing, obtains the compound characteristics of amniotic extracellular matrix dressing of a kind of Bacterial cellulose that contains natural plant composition after sterilizing.
As preferred technical scheme:
Wherein, the preparation method of the compound characteristics of amniotic extracellular matrix dressing of a kind of Bacterial cellulose that contains natural plant composition as above, described can refer to one or more in acetobacter xylinum, rhizobium, Sarcina, Rhodopseudomonas, achromobacter, Alcaligenes, Aerobacter or azotobacter by the cellulosic bacterial strain of secreting bacteria.
The preparation method of the compound characteristics of amniotic extracellular matrix dressing of a kind of Bacterial cellulose that contains natural plant composition as above, the described fermentation medium that contains natural Herba Rosmarini Officinalis lyophilized powder refers in the ethanol water of every 1L and contains natural Herba Rosmarini Officinalis lyophilized powder 10～50g, sodium hydrogen phosphate 4～16g, potassium dihydrogen phosphate 4～16g, yeast extract 4～20g, peptone 4～20g, and wherein ethanol water concentration is 50～70wt%.
The preparation method of the compound characteristics of amniotic extracellular matrix dressing of a kind of Bacterial cellulose that contains natural plant composition as above, described standing cultivation refers to that seed mash mixes with the fermentation medium that contains natural Herba Rosmarini Officinalis lyophilized powder to be placed in culture vessel to leave standstill and cultivates 5～7d, wherein seed mash mix with the fermentation medium that contains collagen solution after liquid level in culture vessel be 0.5～3cm.Under normal circumstances, bacteria cellulose film floats on the top of culture fluid, and Bacterial cellulose can consume a part of culture medium in fermentation culture process, and a part of culture medium loses by evaporation simultaneously.Method described in this patent is by controlling culture fluid consumption at zone of reasonableness, and culture medium consumption is crossed major general affects the biosynthesis of Bacterial cellulose, and affecting at most characteristics of amniotic extracellular matrix material is effectively laid in bacteria cellulose film upper surface excessively.
The preparation method of the compound characteristics of amniotic extracellular matrix dressing of a kind of Bacterial cellulose that contains natural plant composition as above, the described bacteria cellulose film that contains natural plant composition refers to leave standstill cultivates the hydrogel bacteria cellulose film obtaining, thickness is 1～3cm, wherein on the three-dimensional network nanofiber of Bacterial cellulose, is evenly attached with the chemical substance containing natural Herba Rosmarini Officinalis extract composition.First a large amount of amplifications biosynthesis cellulose fento in culture medium of antibacterial in sweat, the chemical substance that contains natural Herba Rosmarini Officinalis extract composition is attached on cellulose fento, along with fermentation time prolongation formation is cellulose nano-fibrous, and on nanofiber, evenly adhere to Herba Rosmarini Officinalis extract composition; The final gluey bacteria cellulose film of the water-setting that contains natural Herba Rosmarini Officinalis extract composition that forms.
The preparation method of the compound characteristics of amniotic extracellular matrix dressing of a kind of Bacterial cellulose that contains natural plant composition as above, described characteristics of amniotic extracellular matrix material refers to source safety, the de-cell biological amniotic membrane of treated removal cell, fat and soluble antigen and dry sterilization.Fresh amnion is taken from health and is cutd open palace and produce puerpera's Placenta Hominis, and antenatal Serological testing is got rid of hepatitis B, hepatitis C, syphilis and adaptive immune deficit syndrome, draws materials to process all under sterile working, to complete; Treated removal cell, fat, soluble antigen, generally adopt freeze-drying to be dried and through irradiation sterilization.
The preparation method of the compound characteristics of amniotic extracellular matrix dressing of a kind of Bacterial cellulose that contains natural plant composition as above, the bacteria cellulose film upper surface that described being laid on contains natural plant composition refers to that the spongy layer of characteristics of amniotic extracellular matrix material contacts with the bacteria cellulose film upper surface that contains natural plant composition.The characteristics of amniotic extracellular matrix material tool structure that is of five storeys can be divided into: epithelial layer, basement membrane layer, compacted zone, fibroblast layer, spongy layer.Wherein the collagen fiber of compacted zone and spongy layer form loose 3 D stereo configuration, are conducive to the network structure that cellulose nano-fibrous and its formation in bacteria cellulose film runs through mutually.Basement membrane layer contains IV type and V Collagen Type VI, laminin，LN, the various chelating albumen of fibronectin Pseudobulbus Bletillae (Rhizoma Bletillae), can promote differentiation, propagation and the migration of seed cell.Therefore, in the time preparing dressing, the spongy layer of characteristics of amniotic extracellular matrix material is contacted with bacteria cellulose film upper surface, be conducive to the cellulose nano-fibrous loose collagen fiber inside that enters spongy layer of bacteria cellulose film, kept the biological activity of basement membrane layer simultaneously.
The preparation method of the compound characteristics of amniotic extracellular matrix dressing of a kind of Bacterial cellulose that contains natural plant composition as above, described purification process refers to composite 30～60 ℃ of immersion 12～24h of sodium hydrate aqueous solution at 1～3wt%, and extremely neutral by washed with de-ionized water.In this patent, select the sodium hydroxide of low concentration at low temperatures composite to be processed, sodium hydroxide solution soaks can thoroughly be removed tropina and stick to the residual media on bacteria cellulose film, guarantee that the endotoxin of composite meets the requirement of medical implanting material, low-concentration sodium hydroxide can prevent from affecting in processing procedure all polyproteins and the somatomedin that in characteristics of amniotic extracellular matrix material, contain simultaneously.
The preparation method of the compound characteristics of amniotic extracellular matrix dressing of a kind of Bacterial cellulose that contains natural plant composition as above, the compound characteristics of amniotic extracellular matrix dressing of the described Bacterial cellulose that contains natural plant composition refers to the bacteria cellulose film that contains natural plant composition and the composite of characteristics of amniotic extracellular matrix, wherein characteristics of amniotic extracellular matrix is positioned at the upper strata of dressing, the bacteria cellulose film that contains natural plant composition is positioned at the lower floor of dressing, and the collagen fiber in compacted zone and the spongy layer of characteristics of amniotic extracellular matrix material run through mutually with the nanofiber in the bacteria cellulose film that contains natural plant composition, formation links composite closely.
The preparation method of the compound characteristics of amniotic extracellular matrix dressing of a kind of Bacterial cellulose that contains natural plant composition as above, in use, the characteristics of amniotic extracellular matrix that is positioned at dressing upper strata contacts with skin wound in the described compound characteristics of amniotic extracellular matrix dressing of a kind of Bacterial cellulose that contains natural plant composition.
Another object of the present invention is to provide the compound characteristics of amniotic extracellular matrix dressing of a kind of Bacterial cellulose that contains natural plant composition, it is characterized in that: comprise Bacterial cellulose, Herba Rosmarini Officinalis and characteristics of amniotic extracellular matrix.
Compared with prior art, the invention has the beneficial effects as follows:
This patent adopts the compound characteristics of amniotic extracellular matrix material of microorganism fermentation culture method original position.The bacteria cellulose material that contains natural Herba Rosmarini Officinalis extract composition and characteristics of amniotic extracellular matrix material, collagen fiber interaction and nanofiber by Bacterial cellulose nanofiber and characteristics of amniotic extracellular matrix material compacted zone and spongy layer run through mutually, form and link composite closely.Guaranteeing, under the histocompatibility of composite, bioactive prerequisite, effectively to have strengthened the mechanical performance of material, anti-microbial property; Contained natural Herba Rosmarini Officinalis extract composition can reduce wound inflammation, strengthen wound contraction, contribute to the reconstruction of epithelial tissue and granulation tissue, and the formation that can improve wound vascularization and collagen protein.Preparation is simple in the present invention, cost is low, material source is extensive, the dressing of preparation has the performances such as sterilization, moisturizing, absorbing wound exudate and accelerating wound healing, is applicable to all kinds of chronic skin wounds such as diabetic foot ulcer, venous leg ulcers, pressure ulcer, skin donor site wound.
The specific embodiment
Below in conjunction with specific embodiment, further set forth the present invention.Should be understood that these embodiment are only not used in and limit the scope of the invention for the present invention is described.In addition should be understood that those skilled in the art can make various changes or modifications the present invention after having read the content of the present invention's instruction, these equivalent form of values fall within the application's appended claims limited range equally.
Embodiment 1:
Choose and can the cellulosic acetobacter xylinum activation of secreting bacteria be prepared into seed mash, the seed mash that is then 50wt% by bacterial strain concentration mixes with the fermentation medium that contains natural Herba Rosmarini Officinalis lyophilized powder.The described fermentation medium that contains natural Herba Rosmarini Officinalis lyophilized powder refers in the ethanol water of every 1L and contains natural Herba Rosmarini Officinalis lyophilized powder 30g, sodium hydrogen phosphate 16g, potassium dihydrogen phosphate 16g, yeast extract 20g, peptone 20g, and wherein ethanol water concentration is 70wt%.Mixed solution is placed in culture vessel, and the liquid level of mixed liquor in culture vessel is 3cm, leaves standstill to cultivate 5d to obtain film thickness be the bacteria cellulose film that the water-setting glue of 3cm contains natural Herba Rosmarini Officinalis extract composition.
The safety of originating, the characteristics of amniotic extracellular matrix material of treated removal cell, fat and soluble antigen and dry sterilization is immersed in culture medium, is laid in bacteria cellulose film upper surface after its complete wetting.Wherein, the spongy layer of characteristics of amniotic extracellular matrix material contacts with the bacteria cellulose film upper surface that contains natural plant composition.Continue to leave standstill and cultivate 1h, the composite obtaining is placed in 60 ℃ of immersion 12h of sodium hydrate aqueous solution of 1wt%, and extremely neutral by washed with de-ionized water, obtains the compound characteristics of amniotic extracellular matrix dressing of a kind of Bacterial cellulose that contains natural plant composition.Wherein characteristics of amniotic extracellular matrix is positioned at the upper strata of dressing, the bacteria cellulose film that contains natural plant composition is positioned at the lower floor of dressing, and the collagen fiber in compacted zone and the spongy layer of characteristics of amniotic extracellular matrix material run through mutually with the nanofiber in the bacteria cellulose film that contains natural plant composition, form and link composite closely.In use, the characteristics of amniotic extracellular matrix that is positioned at dressing upper strata contacts with skin wound in this dressing.
Embodiment 2:
Choose can the cellulosic rhizobium of secreting bacteria and Rhodopseudomonas activation be prepared into seed mash, the seed mash that is then 40wt% by bacterial strain concentration mixes with the fermentation medium that contains natural Herba Rosmarini Officinalis lyophilized powder.The described fermentation medium that contains natural Herba Rosmarini Officinalis lyophilized powder refers in the ethanol water of every 1L and contains natural Herba Rosmarini Officinalis lyophilized powder 20g, sodium hydrogen phosphate 14g, potassium dihydrogen phosphate 14g, yeast extract 18g, peptone 18g, and wherein ethanol water concentration is 70wt%.Mixed solution is placed in culture vessel, and the liquid level of mixed liquor in culture vessel is 2cm, leaves standstill to cultivate 5d to obtain film thickness be the bacteria cellulose film that the water-setting glue of 2.5cm contains natural Herba Rosmarini Officinalis extract composition.
The safety of originating, the characteristics of amniotic extracellular matrix material of treated removal cell, fat and soluble antigen and dry sterilization is immersed in culture medium, is laid in bacteria cellulose film upper surface after its complete wetting.Wherein, the spongy layer of characteristics of amniotic extracellular matrix material contacts with the bacteria cellulose film upper surface that contains natural plant composition.Continue to leave standstill and cultivate 2h, the composite obtaining is placed in 50 ℃ of immersion 12h of sodium hydrate aqueous solution of 2wt%, and extremely neutral by washed with de-ionized water, obtains the compound characteristics of amniotic extracellular matrix dressing of a kind of Bacterial cellulose that contains natural plant composition.Wherein characteristics of amniotic extracellular matrix is positioned at the upper strata of dressing, the bacteria cellulose film that contains natural plant composition is positioned at the lower floor of dressing, and the collagen fiber in compacted zone and the spongy layer of characteristics of amniotic extracellular matrix material run through mutually with the nanofiber in the bacteria cellulose film that contains natural plant composition, form and link composite closely.In use, the characteristics of amniotic extracellular matrix that is positioned at dressing upper strata contacts with skin wound in this dressing.
Embodiment 3:
Choose can the cellulosic Sarcina of secreting bacteria and Alcaligenes activation be prepared into seed mash, the seed mash that is then 30wt% by bacterial strain concentration mixes with the fermentation medium that contains natural Herba Rosmarini Officinalis lyophilized powder.The described fermentation medium that contains natural Herba Rosmarini Officinalis lyophilized powder refers in the ethanol water of every 1L and contains natural Herba Rosmarini Officinalis lyophilized powder 10g, sodium hydrogen phosphate 12g, potassium dihydrogen phosphate 12g, yeast extract 16g, peptone 16g, and wherein ethanol water concentration is 60wt%.Mixed solution is placed in culture vessel, and the liquid level of mixed liquor in culture vessel is 1cm, leaves standstill to cultivate 6d to obtain film thickness be the bacteria cellulose film that the water-setting glue of 1cm contains natural Herba Rosmarini Officinalis extract composition.
The safety of originating, the characteristics of amniotic extracellular matrix material of treated removal cell, fat and soluble antigen and dry sterilization is immersed in culture medium, is laid in bacteria cellulose film upper surface after its complete wetting.Wherein, the spongy layer of characteristics of amniotic extracellular matrix material contacts with the bacteria cellulose film upper surface that contains natural plant composition.Continue to leave standstill and cultivate 3h, the composite obtaining is placed in 30 ℃ of immersion 16h of sodium hydrate aqueous solution of 3wt%, and extremely neutral by washed with de-ionized water, obtains the compound characteristics of amniotic extracellular matrix dressing of a kind of Bacterial cellulose that contains natural plant composition.Wherein characteristics of amniotic extracellular matrix is positioned at the upper strata of dressing, the bacteria cellulose film that contains natural plant composition is positioned at the lower floor of dressing, and the collagen fiber in compacted zone and the spongy layer of characteristics of amniotic extracellular matrix material run through mutually with the nanofiber in the bacteria cellulose film that contains natural plant composition, form and link composite closely.In use, the characteristics of amniotic extracellular matrix that is positioned at dressing upper strata contacts with skin wound in this dressing.
Embodiment 4:
Choose the cellulosic acetobacter xylinum of energy secreting bacteria, Aerobacter and azotobacter activation and be prepared into seed mash, the seed mash that is then 50wt% by bacterial strain concentration mixes with the fermentation medium that contains natural Herba Rosmarini Officinalis lyophilized powder.The described fermentation medium that contains natural Herba Rosmarini Officinalis lyophilized powder refers in the ethanol water of every 1L and contains natural Herba Rosmarini Officinalis lyophilized powder 50g, sodium hydrogen phosphate 10g, potassium dihydrogen phosphate 10g, yeast extract 10g, peptone 10g, and wherein ethanol water concentration is 60wt%.Mixed solution is placed in culture vessel, and the liquid level of mixed liquor in culture vessel is 0.5cm, leaves standstill to cultivate 6d to obtain film thickness be the bacteria cellulose film that the water-setting glue of 2cm contains natural Herba Rosmarini Officinalis extract composition.
The safety of originating, the characteristics of amniotic extracellular matrix material of treated removal cell, fat and soluble antigen and dry sterilization is immersed in culture medium, is laid in bacteria cellulose film upper surface after its complete wetting.Wherein, the spongy layer of characteristics of amniotic extracellular matrix material contacts with the bacteria cellulose film upper surface that contains natural plant composition.Continue to leave standstill and cultivate 3h, the composite obtaining is placed in 30 ℃ of immersion 24h of sodium hydrate aqueous solution of 1wt%, and extremely neutral by washed with de-ionized water, obtains the compound characteristics of amniotic extracellular matrix dressing of a kind of Bacterial cellulose that contains natural plant composition.Wherein characteristics of amniotic extracellular matrix is positioned at the upper strata of dressing, the bacteria cellulose film that contains natural plant composition is positioned at the lower floor of dressing, and the collagen fiber in compacted zone and the spongy layer of characteristics of amniotic extracellular matrix material run through mutually with the nanofiber in the bacteria cellulose film that contains natural plant composition, form and link composite closely.In use, the characteristics of amniotic extracellular matrix that is positioned at dressing upper strata contacts with skin wound in this dressing.
Embodiment 5:
Choose can the cellulosic acetobacter xylinum of secreting bacteria and achromobacter activation be prepared into seed mash, the seed mash that is then 40wt% by bacterial strain concentration mixes with the fermentation medium that contains natural Herba Rosmarini Officinalis lyophilized powder.The described fermentation medium that contains natural Herba Rosmarini Officinalis lyophilized powder refers in the ethanol water of every 1L and contains natural Herba Rosmarini Officinalis lyophilized powder 40g, sodium hydrogen phosphate 4g, potassium dihydrogen phosphate 4g, yeast extract 4g, peptone 4g, and wherein ethanol water concentration is 50wt%.Mixed solution is placed in culture vessel, and the liquid level of mixed liquor in culture vessel is 2cm, leaves standstill to cultivate 7d to obtain film thickness be the bacteria cellulose film that the water-setting glue of 2cm contains natural Herba Rosmarini Officinalis extract composition.
The safety of originating, the characteristics of amniotic extracellular matrix material of treated removal cell, fat and soluble antigen and dry sterilization is immersed in culture medium, is laid in bacteria cellulose film upper surface after its complete wetting.Wherein, the spongy layer of characteristics of amniotic extracellular matrix material contacts with the bacteria cellulose film upper surface that contains natural plant composition.Continue to leave standstill and cultivate 6h, the composite obtaining is placed in 40 ℃ of immersion 18h of sodium hydrate aqueous solution of 3wt%, and extremely neutral by washed with de-ionized water, obtains the compound characteristics of amniotic extracellular matrix dressing of a kind of Bacterial cellulose that contains natural plant composition.Wherein characteristics of amniotic extracellular matrix is positioned at the upper strata of dressing, the bacteria cellulose film that contains natural plant composition is positioned at the lower floor of dressing, and the collagen fiber in compacted zone and the spongy layer of characteristics of amniotic extracellular matrix material run through mutually with the nanofiber in the bacteria cellulose film that contains natural plant composition, form and link composite closely.In use, the characteristics of amniotic extracellular matrix that is positioned at dressing upper strata contacts with skin wound in this dressing.
Embodiment 6:
The subcutaneous implantation experiment of the compound characteristics of amniotic extracellular matrix dressing of Bacterial cellulose that contains natural plant composition
Experimental technique: get 30 of healthy SD rats, male or female, is divided into experimental group and matched group, sterilization, adopts ether inhalation anesthesia, and skin of back cuts in conventional sterile working, and 51 × 1cm are implanted at experimental group back
2dressing (getting respectively the compound characteristics of amniotic extracellular matrix dressing of the prepared Bacterial cellulose that contains natural plant composition of embodiment 1-5), matched group is sewed up after cutting.Observe the postoperative ordinary circumstance of animal and wound healing situation.Within postoperative 1,2,4 week, put to death each treated animal, take out implantation dressing glutaraldehyde and fix, conventional embedded section, HE dyeing, om observation implant peripheral lymphocyte, macrophages infiltration situation.
Experiment shows: after operation animal on the same day is clear-headed, can normally take food, wound is without inflammatory reaction such as redness, sepages, and all wound surface all can well heal, and operative incision infects without 1 example, animal is without toxic reactions such as death, convulsions, apoplexy, implant surrounding soft tissue has no the performances such as blackout, necrosis, suppuration, hydrops, postoperative 1 week of histological observation, and implant has large amount lymphocyte around, 2 weeks time, only a small amount of lymphocyte, 4 weeks time, exists without lymphocyte substantially.Confirm dressing abiology toxicity of the present invention, have good histocompatibility.
Embodiment 7
The treatment experiment of the compound characteristics of amniotic extracellular matrix dressing of Bacterial cellulose that contains natural plant composition to diabetic foot ulcer rat
Use high lipid food to coordinate the rats with type 2 diabetes of streptozotocin induction, use scald apparatus to make dark II degree scald induction skin ulcer and make diabetic foot ulcer rat model.Rat is SD male rat, SPF level, body weight (180 ± 10) g, totally 30,5 every group, totally 6 groups, wherein one group is contrast, uses respectively the compound characteristics of amniotic extracellular matrix dressing of the Bacterial cellulose that contains natural plant composition prepared by embodiment 1-5 for another 5 groups.The preparation method of high lipid food is: first, by Fructus Hordei Vulgaris 15%, Semen Tritici aestivi 31%, Semen Maydis 15%, wheat bran 15%, after fish flour 6% is uniformly mixed in proportion, add bean cake, oil, salt to mix rear extrusion forming and make normal feedstuff, then, by normal feedstuff 73%, Adeps Sus domestica 20%, milk powder 2%, cholesterol 1%, sucrose 4% is uniformly mixed in proportion rear extrusion forming and makes high lipid food, and finished product is cylinder bulk.High lipid food was fed after 8 weeks, and 1.5% streptozotocin is dissolved in the citric acid-sodium citrate buffer of pH value 4.5 by the dosage left lower quadrant injection of 35mg/kg.Use SQL-5Q scald apparatus, 90 ℃ of temperature, time of contact 10S, pressure 9.8N, area 4cm
2, after anesthesia, scald in back part skin.After modeling success, with the clean wound surface of povidone iodine, with the compound characteristics of amniotic extracellular matrix dressing of the Bacterial cellulose that contains natural plant composition of the present invention external application wound surface, then wrap up with sterile gauze, within 1 day, change dressings 1 time, add up to indicator-specific statistics the average healing with ulcer area and blood leukocytes, the results are shown in Table 1.
The average healing comparison (n=5) of table 1 to diabetic foot ulcer rat
| ? | Embodiment 1 | Embodiment 2 | Embodiment 3 | Embodiment 4 | Embodiment 5 |
| The average healing/sky | 18.5±1.4 | 17.8±0.8 | 18.4±1.2 | 19.2±1.0 | 17.0±1.2 |
Embodiment 8:
The hospital clinical trial of the compound characteristics of amniotic extracellular matrix dressing of Bacterial cellulose that contains natural plant composition
The compound characteristics of amniotic extracellular matrix dressing of Bacterial cellulose that prepared by the embodiment of the present invention 1 contain natural plant composition is through hospital clinical trial treatment totally 102 examples.Wherein relate to diabetic foot ulcer 37 examples, venous leg ulcers 21 examples, skin infection or carbuncle furuncle 44 examples, its therapeutic effect refers to table 2.
In table 2, the curative effect determinate standard of various diabetic foot ulcers, venous leg ulcers, skin infection or carbuncle furuncle is:
1. recovery from illness: General Symptoms disappears, local swelling is dissipated, or pus absorption dissipation, or skin ulcer face healing after bursting, and blood leukocytes is normally total.
2. effective: General Symptoms disappears, and swelling, abscess or the open sore part have been dwindled more than 70%, blood leukocytes sum is normal.
3. effective: General Symptoms alleviates, swelling, abscess or the open sore part have been dwindled more than 30%, and below 70%, blood leukocytes sum approaches normal.
4. invalid: not reach effective standard.
The clinical experiment of the compound characteristics of amniotic extracellular matrix dressing of Bacterial cellulose that table 2 contains natural plant composition
| Case type | Case load | Recovery from illness | Effective | Effectively | Invalid | Cure rate |
| Diabetic foot ulcer | 37 | 36 | 1 | 0 | 0 | 97.3% |
| Venous leg ulcers | 21 | 19 | 1 | 1 | 0 | 90.5% |
| Skin infection or carbuncle furuncle | 44 | 42 | 1 | 1 | 0 | 95.5% |
Wherein concrete case has:
Week certain, man, 68 years old, diabetic history 20 years, fasting glucose 14 mnol/l, early 2 hours blood glucose 17.9 mmol/L after the meal, glycolated hemoglobin 11.2 mg/dl, three grades of diabetic foot ulcers, red swelling of the skin, fester, deep ulcer, forms ulcer chamber, ulcer area 145.5 mm
2, blood leukocytes sum 1.8 × 10
9/ l, neutrophilic granulocyte number 77%.With the clean wound surface of povidone iodine, remove slough and purulent secretion, clog in intracavity and external application wound surface with the compound characteristics of amniotic extracellular matrix dressing of the Bacterial cellulose that contains natural plant composition of the present invention, then wrap up with sterile gauze, lifting affected limb, does not make foot pressurized, within 1 day, changes dressings 1 time, treat 12 days courses for the treatment of continuously, ulcer area reduces to 64.5 mm
2, blood leukocytes sum is down to 0.7 × 10
9/ l, neutrophilic granulocyte number 55%.Continued treatment to 24 day, muscular tissue layer, fat deposit, skin corium, epidermal area recover original organizational structure, cure completely.
Wang, female, 47 years old, fasting glucose 8.1 mmol/L, early 2 hours blood glucose 10.4 mmol/L after the meal, glycolated hemoglobin 8.7 mg/dl, left sufficient diabetic foot ulcer one-level, ulcer area 123.3 mm
2, suppurating sore, blood leukocytes sum 1.6 × 10
9/ l, neutrophilic granulocyte number 70%.With the clean wound surface of povidone iodine, remove slough and purulent secretion, with the compound characteristics of amniotic extracellular matrix dressing of the Bacterial cellulose that contains natural plant composition of the present invention external application wound surface, then wrap up with sterile gauze, lifting affected limb, does not make foot pressurized, within 1 day, changes dressings 1 time, 7 days local symptoms for the treatment of disappear continuously, and blood leukocytes sum is down to 0.5 × 10
9/ l, neutrophilic granulocyte number 48%, ulcer surface heals completely.
Jiao, female, 67 years old, skin infection, intractable ulcer patient, left lower extremity ulcer 5 wheat harvesting periods, part presents swollen, hard tuberosity, has risen root bundle, and skin ulcer color is red, there is purulent core on skin ulcer top, and pain is obvious, and blood leukocytes sum and neutrophilic granulocyte number increase, blood leukocytes sum 1.4 × 10
9/ l, neutrophilic granulocyte number 73%.With the clean wound surface of povidone iodine, remove slough and purulent secretion, adopt on the compound characteristics of amniotic extracellular matrix dressing of the Bacterial cellulose that contains natural plant composition of the present invention external application ulcer surface, then wrap up with sterile gauze, within 1 day, change dressings 1 time, medication is after 7 days, and redness obviously alleviates, ulcer surface dwindles, and blood leukocytes sum is down to 0.7 × 10
9/ l, neutrophilic granulocyte number 60%.Continue medication to the 14 days, left lower extremity is without red, swollen, and without pain, ulcer wound surface heals, and blood leukocytes sum is down to 0.6 × 10
9/ l, neutrophilic granulocyte number 53%, does not stay obvious cicatrix, treatment recovery from illness.
Claims (11)
1. contain the compound characteristics of amniotic extracellular matrix dressing of Bacterial cellulose of natural plant composition, it is characterized in that: comprise Bacterial cellulose, Herba Rosmarini Officinalis and characteristics of amniotic extracellular matrix.
2. the preparation method of the compound characteristics of amniotic extracellular matrix dressing of the Bacterial cellulose that contains natural plant composition, it is characterized in that: choose can secreting bacteria cellulosic bacterial strain activation and be prepared into seed mash, the seed mash that is then 30～50wt% by bacterial strain concentration mixes with the fermentation medium that contains natural Herba Rosmarini Officinalis lyophilized powder and is placed on standing cultivation in culture vessel and obtains the bacteria cellulose film that contains natural plant composition; Get characteristics of amniotic extracellular matrix film and be immersed in culture medium, after its complete wetting, be laid in bacteria cellulose film upper surface; Continue to leave standstill and cultivate 1～6h, purified processing, obtains the compound characteristics of amniotic extracellular matrix dressing of a kind of Bacterial cellulose that contains natural plant composition after sterilizing.
3. the preparation method of the compound characteristics of amniotic extracellular matrix dressing of a kind of Bacterial cellulose that contains natural plant composition as claimed in claim 2, is characterized in that: described can refer to one or more in acetobacter xylinum, rhizobium, Sarcina, Rhodopseudomonas, achromobacter, Alcaligenes, Aerobacter or azotobacter by the cellulosic bacterial strain of secreting bacteria.
4. the preparation method of the compound characteristics of amniotic extracellular matrix dressing of a kind of Bacterial cellulose that contains natural plant composition as claimed in claim 2, it is characterized in that: the described fermentation medium that contains natural Herba Rosmarini Officinalis lyophilized powder refers in the ethanol water of every 1L and contain natural Herba Rosmarini Officinalis lyophilized powder 10～50g, sodium hydrogen phosphate 4～16g, potassium dihydrogen phosphate 4～16g, yeast extract 4～20g, peptone 4～20g, wherein ethanol water concentration is 50～70wt%.
5. the preparation method of the compound characteristics of amniotic extracellular matrix dressing of a kind of Bacterial cellulose that contains natural plant composition as claimed in claim 2, it is characterized in that: described standing cultivation refers to that seed mash mixes with the fermentation medium that contains natural Herba Rosmarini Officinalis lyophilized powder to be placed in culture vessel to leave standstill and cultivate 5～7d, wherein seed mash mix with the fermentation medium that contains collagen solution after liquid level in culture vessel be 0.5～3cm.
6. the preparation method of the compound characteristics of amniotic extracellular matrix dressing of a kind of Bacterial cellulose that contains natural plant composition as claimed in claim 2, it is characterized in that: the bacteria cellulose film that contains natural plant composition refers to leave standstill cultivates the hydrogel bacteria cellulose film obtaining, thickness is 1～3cm, wherein on the three-dimensional network nanofiber of Bacterial cellulose, is evenly attached with the chemical substance containing natural Herba Rosmarini Officinalis extract composition.
7. the preparation method of the compound characteristics of amniotic extracellular matrix dressing of a kind of Bacterial cellulose that contains natural plant composition as claimed in claim 2, it is characterized in that: described characteristics of amniotic extracellular matrix material refers to source safety the de-cell biological amniotic membrane of treated removal cell, fat and soluble antigen and dry sterilization.
8. the preparation method of the compound characteristics of amniotic extracellular matrix dressing of a kind of Bacterial cellulose that contains natural plant composition as claimed in claim 2, is characterized in that: the bacteria cellulose film upper surface that described being laid in contains natural plant composition refers to that the spongy layer of characteristics of amniotic extracellular matrix material contacts with the bacteria cellulose film upper surface that contains natural plant composition.
9. the preparation method of the compound characteristics of amniotic extracellular matrix dressing of a kind of Bacterial cellulose that contains natural plant composition as claimed in claim 2, it is characterized in that: described purification process refers to composite 30～60 ℃ of immersion 12～24h of sodium hydrate aqueous solution at 1～3wt%, and extremely neutral by washed with de-ionized water.
10. the preparation method of the compound characteristics of amniotic extracellular matrix dressing of a kind of Bacterial cellulose that contains natural plant composition as claimed in claim 2, it is characterized in that: the compound characteristics of amniotic extracellular matrix dressing of the described Bacterial cellulose that contains natural plant composition refers to the bacteria cellulose film that contains natural plant composition and the composite of characteristics of amniotic extracellular matrix, wherein characteristics of amniotic extracellular matrix is positioned at the upper strata of dressing, the bacteria cellulose film that contains natural plant composition is positioned at the lower floor of dressing, and the collagen fiber in compacted zone and the spongy layer of characteristics of amniotic extracellular matrix material run through mutually with the nanofiber in the bacteria cellulose film that contains natural plant composition, formation links composite closely.
The preparation method of the compound characteristics of amniotic extracellular matrix dressing of 11. a kind of Bacterial cellulose that contains natural plant composition as claimed in claim 2, it is characterized in that: in use, the characteristics of amniotic extracellular matrix that is positioned at dressing upper strata contacts with skin wound in the described compound characteristics of amniotic extracellular matrix dressing of a kind of Bacterial cellulose that contains natural plant composition.
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| CN111481735A (en) * | 2019-01-25 | 2020-08-04 | 华中科技大学同济医学院附属协和医院 | Medical antibacterial wound-protecting hydrogel dressing and preparation method thereof |
| CN111790000A (en) * | 2020-06-23 | 2020-10-20 | 洛阳巴库生物科技有限公司 | Stem cell amniotic membrane dressing structure for wound repair and preparation method thereof |
| CN116802228A (en) * | 2021-02-24 | 2023-09-22 | 海南光宇生物科技有限公司 | Bacterial cellulose-based porous foam dressing, and preparation method and application thereof |
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| CN116802228A (en) * | 2021-02-24 | 2023-09-22 | 海南光宇生物科技有限公司 | Bacterial cellulose-based porous foam dressing, and preparation method and application thereof |
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| CN103893816B (en) | 2015-06-10 |
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