CN113440523B - Chitosan-modified benzisothiazolinone polylactic acid nanosphere and preparation method and application thereof - Google Patents
Chitosan-modified benzisothiazolinone polylactic acid nanosphere and preparation method and application thereof Download PDFInfo
- Publication number
- CN113440523B CN113440523B CN202110619426.5A CN202110619426A CN113440523B CN 113440523 B CN113440523 B CN 113440523B CN 202110619426 A CN202110619426 A CN 202110619426A CN 113440523 B CN113440523 B CN 113440523B
- Authority
- CN
- China
- Prior art keywords
- chitosan
- nanospheres
- bit
- benzisothiazolinone
- pla
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Active
Links
Images
Classifications
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K47/00—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient
- A61K47/50—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates
- A61K47/69—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates the conjugate being characterised by physical or galenical forms, e.g. emulsion, particle, inclusion complex, stent or kit
- A61K47/6921—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates the conjugate being characterised by physical or galenical forms, e.g. emulsion, particle, inclusion complex, stent or kit the form being a particulate, a powder, an adsorbate, a bead or a sphere
- A61K47/6927—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates the conjugate being characterised by physical or galenical forms, e.g. emulsion, particle, inclusion complex, stent or kit the form being a particulate, a powder, an adsorbate, a bead or a sphere the form being a solid microparticle having no hollow or gas-filled cores
- A61K47/6929—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates the conjugate being characterised by physical or galenical forms, e.g. emulsion, particle, inclusion complex, stent or kit the form being a particulate, a powder, an adsorbate, a bead or a sphere the form being a solid microparticle having no hollow or gas-filled cores the form being a nanoparticle, e.g. an immuno-nanoparticle
- A61K47/6931—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates the conjugate being characterised by physical or galenical forms, e.g. emulsion, particle, inclusion complex, stent or kit the form being a particulate, a powder, an adsorbate, a bead or a sphere the form being a solid microparticle having no hollow or gas-filled cores the form being a nanoparticle, e.g. an immuno-nanoparticle the material constituting the nanoparticle being a polymer
- A61K47/6935—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates the conjugate being characterised by physical or galenical forms, e.g. emulsion, particle, inclusion complex, stent or kit the form being a particulate, a powder, an adsorbate, a bead or a sphere the form being a solid microparticle having no hollow or gas-filled cores the form being a nanoparticle, e.g. an immuno-nanoparticle the material constituting the nanoparticle being a polymer the polymer being obtained otherwise than by reactions involving carbon to carbon unsaturated bonds, e.g. polyesters, polyamides or polyglycerol
- A61K47/6937—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates the conjugate being characterised by physical or galenical forms, e.g. emulsion, particle, inclusion complex, stent or kit the form being a particulate, a powder, an adsorbate, a bead or a sphere the form being a solid microparticle having no hollow or gas-filled cores the form being a nanoparticle, e.g. an immuno-nanoparticle the material constituting the nanoparticle being a polymer the polymer being obtained otherwise than by reactions involving carbon to carbon unsaturated bonds, e.g. polyesters, polyamides or polyglycerol the polymer being PLGA, PLA or polyglycolic acid
-
- A—HUMAN NECESSITIES
- A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
- A01N—PRESERVATION OF BODIES OF HUMANS OR ANIMALS OR PLANTS OR PARTS THEREOF; BIOCIDES, e.g. AS DISINFECTANTS, AS PESTICIDES OR AS HERBICIDES; PEST REPELLANTS OR ATTRACTANTS; PLANT GROWTH REGULATORS
- A01N25/00—Biocides, pest repellants or attractants, or plant growth regulators, characterised by their forms, or by their non-active ingredients or by their methods of application, e.g. seed treatment or sequential application; Substances for reducing the noxious effect of the active ingredients to organisms other than pests
- A01N25/08—Biocides, pest repellants or attractants, or plant growth regulators, characterised by their forms, or by their non-active ingredients or by their methods of application, e.g. seed treatment or sequential application; Substances for reducing the noxious effect of the active ingredients to organisms other than pests containing solids as carriers or diluents
- A01N25/10—Macromolecular compounds
-
- A—HUMAN NECESSITIES
- A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
- A01N—PRESERVATION OF BODIES OF HUMANS OR ANIMALS OR PLANTS OR PARTS THEREOF; BIOCIDES, e.g. AS DISINFECTANTS, AS PESTICIDES OR AS HERBICIDES; PEST REPELLANTS OR ATTRACTANTS; PLANT GROWTH REGULATORS
- A01N43/00—Biocides, pest repellants or attractants, or plant growth regulators containing heterocyclic compounds
- A01N43/72—Biocides, pest repellants or attractants, or plant growth regulators containing heterocyclic compounds having rings with nitrogen atoms and oxygen or sulfur atoms as ring hetero atoms
- A01N43/80—Biocides, pest repellants or attractants, or plant growth regulators containing heterocyclic compounds having rings with nitrogen atoms and oxygen or sulfur atoms as ring hetero atoms five-membered rings with one nitrogen atom and either one oxygen atom or one sulfur atom in positions 1,2
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K31/00—Medicinal preparations containing organic active ingredients
- A61K31/33—Heterocyclic compounds
- A61K31/395—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins
- A61K31/41—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins having five-membered rings with two or more ring hetero atoms, at least one of which being nitrogen, e.g. tetrazole
- A61K31/425—Thiazoles
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K47/00—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient
- A61K47/50—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates
- A61K47/51—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates the non-active ingredient being a modifying agent
- A61K47/56—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates the non-active ingredient being a modifying agent the modifying agent being an organic macromolecular compound, e.g. an oligomeric, polymeric or dendrimeric molecule
- A61K47/61—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates the non-active ingredient being a modifying agent the modifying agent being an organic macromolecular compound, e.g. an oligomeric, polymeric or dendrimeric molecule the organic macromolecular compound being a polysaccharide or a derivative thereof
-
- 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
-
- 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
- A61P31/10—Antimycotics
-
- B—PERFORMING OPERATIONS; TRANSPORTING
- B82—NANOTECHNOLOGY
- B82Y—SPECIFIC USES OR APPLICATIONS OF NANOSTRUCTURES; MEASUREMENT OR ANALYSIS OF NANOSTRUCTURES; MANUFACTURE OR TREATMENT OF NANOSTRUCTURES
- B82Y30/00—Nanotechnology for materials or surface science, e.g. nanocomposites
-
- B—PERFORMING OPERATIONS; TRANSPORTING
- B82—NANOTECHNOLOGY
- B82Y—SPECIFIC USES OR APPLICATIONS OF NANOSTRUCTURES; MEASUREMENT OR ANALYSIS OF NANOSTRUCTURES; MANUFACTURE OR TREATMENT OF NANOSTRUCTURES
- B82Y40/00—Manufacture or treatment of nanostructures
-
- B—PERFORMING OPERATIONS; TRANSPORTING
- B82—NANOTECHNOLOGY
- B82Y—SPECIFIC USES OR APPLICATIONS OF NANOSTRUCTURES; MEASUREMENT OR ANALYSIS OF NANOSTRUCTURES; MANUFACTURE OR TREATMENT OF NANOSTRUCTURES
- B82Y5/00—Nanobiotechnology or nanomedicine, e.g. protein engineering or drug delivery
-
- Y—GENERAL TAGGING OF NEW TECHNOLOGICAL DEVELOPMENTS; GENERAL TAGGING OF CROSS-SECTIONAL TECHNOLOGIES SPANNING OVER SEVERAL SECTIONS OF THE IPC; TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
- Y02—TECHNOLOGIES OR APPLICATIONS FOR MITIGATION OR ADAPTATION AGAINST CLIMATE CHANGE
- Y02A—TECHNOLOGIES FOR ADAPTATION TO CLIMATE CHANGE
- Y02A50/00—TECHNOLOGIES FOR ADAPTATION TO CLIMATE CHANGE in human health protection, e.g. against extreme weather
- Y02A50/30—Against vector-borne diseases, e.g. mosquito-borne, fly-borne, tick-borne or waterborne diseases whose impact is exacerbated by climate change
Abstract
The invention discloses a chitosan-modified benzisothiazolinone polylactic acid nanosphere as well as a preparation method and application thereof. Benzo-isothiazolinone is loaded on polylactic acid nanospheres, and then the benzo-isothiazolinone-loaded polylactic acid nanospheres are modified by chitosan to obtain chitosan-modified benzo-isothiazolinone polylactic acid nanospheres. According to the invention, a solvent volatilization method is adopted to prepare the PLA nanosphere loaded with the BIT antibacterial drug, and chitosan is modified on the surface of the BIT-PLA nanosphere to obtain the chitosan-modified benzisothiazolinone polylactic acid nanosphere. The prepared chitosan-modified benzisothiazolinone polylactic acid nanospheres can effectively improve the transmembrane transport rate and antibacterial activity of a nano drug-loaded system, thereby being beneficial to the efficient utilization of benzisothiazolinone in the antibacterial fields of mildew prevention, corrosion prevention and the like and overcoming the problems of insolubility, dispersibility and stability of benzisothiazolinone.
Description
Technical Field
The invention belongs to the field of antibacterial materials, and particularly relates to chitosan-modified benzisothiazolinone polylactic acid nanospheres as well as a preparation method and application thereof.
Background
Benzisothiazolinone (1, 2-benzisothiazolin-3-one, BIT) and derivative compounds thereof have penetrability on target receptor cell membranes, and the BIT interacts with the cell membranes and intracellular sulfhydryl proteins (glutathione, cysteine, thioglycolic acid, mercaptoethanol and the like) through transmembrane transport to break S-N bonds of the sulfhydryl-containing proteins, so that the S-N bonds are combined with corresponding receptors to form S-S bonds, protein denaturation inactivation is caused, the cell membrane structure is damaged, the normal functions of cells are damaged, apoptosis is finally caused, and the growth and propagation of harmful microorganisms such as bacteria, fungi and the like are effectively inhibited. Therefore, BIT has the excellent performances of wide antimicrobial spectrum, environmental safety and the like, is widely applied to industrial antibiosis, mold prevention and marine antifouling, and has made great research progress in the fields of agriculture and medicine. The problems of insolubility, dispersibility and stability of BIT seriously restrict the high-efficiency utilization of BIT in the antibacterial fields of mildew prevention, corrosion prevention and the like.
The nano drug-carrying system is a novel cross-assembly group fused with multiple disciplines such as materials science, physical chemistry, biomedicine and the like, and has the advantages of long acting, sustained and controlled release, targeted delivery and the like. Has great development space in the fields of disease prevention and control, medical development, antibiosis and the like. Polylactic acid (PLA) artificially synthesized biodegradable polyester-based polymer materials have been approved by the U.S. food and drug administration and the european drug administration for use in the medical field. PLA is hydrolyzed into a lactic acid monomer through ester bonds, and is circularly metabolized through citric acid to generate nontoxic carbon dioxide and water. PLA is used as a drug carrier, has the advantages of high mechanical strength, good biocompatibility and the like, can effectively improve drug absorption, prolong drug duration and further improve bioavailability.
Chitosan (CS) is a natural cationic polysaccharide, is derived from shells of crustaceans by deacetylation of chitin under alkaline conditions, has the yield second to that of the second largest natural high-molecular polysaccharide of cellulose, has excellent biocompatibility, biodegradability and antibacterial performance, and has wide application prospect in the field of biomedicine, particularly in the aspect of interface modification of drug carriers.
Disclosure of Invention
The invention aims to overcome the defect of low antibacterial activity of insoluble BIT (built-in kit) medicaments, and provides chitosan-modified benzisothiazolinone polylactic acid nanospheres which can enhance the transmembrane transport rate of medicaments in thalli so as to improve the pair effect of medicinal preparationsE. coliAndS. aureusand (4) antibacterial activity.
The invention discloses a chitosan-modified benzisothiazolinone polylactic acid nanosphere, which is prepared by loading benzisothiazolinone on a polylactic acid nanosphere, and then modifying the benzisothiazolinone-loaded polylactic acid nanosphere with chitosan.
Preferably, the average particle size of the chitosan modified benzisothiazolinone polylactic acid nanosphere is 186.47-206.57 nm, and the corresponding zeta potential is 0.76-25.61 mV.
Preferably, the specific preparation method comprises the following steps:
dissolving polylactic acid and benzisothiazolinone in dichloromethane to prepare an organic phase, dissolving polyvinyl alcohol in deionized water to prepare a water phase, adding the organic phase in the water phase, uniformly stirring, ultrasonically emulsifying, stirring and solidifying emulsion, centrifugally collecting solid from the solidified BIT-PLA nanospheres, washing the solid with deionized water, freeze-drying to prepare BIT-PLA nanospheres, dispersing the BIT-PLA nanospheres in chitosan solution, stirring and reacting to modify the BIT-PLA nanospheres with chitosan, collecting the solid, and washing the solid with the deionized water to prepare the benzisothiazolinone polylactic acid nanospheres modified by the chitosan.
Preferably, the dosage mass ratio of the polylactic acid, the benzisothiazolinone and the chitosan is as follows: 70-90:20:20-100. More preferably 80:20:20-100.
Preferably, the ultrasonic emulsification is ultrasonic emulsification for 3 minutes by a probe type ultrasonicator under the ice bath condition of 360 w for 3-7 s and 5 s in the gap.
Preferably, the stirring solidification of the emulsion is to stir and solidify the emulsion at the room temperature at 700-900rpm for 10-14 hours.
Preferably, the solid is collected by centrifuging the solidified BIT-PLA nanospheres at 8000-12000 rpm at 22-28 ℃ for 25-35 minutes.
Preferably, 70-90mg of polylactic acid and 20 mg of benzisothiazolinone are dissolved in 5 mL of dichloromethane to prepare an organic phase, 120 mg of polyvinyl alcohol is dissolved in 18-22 mL of deionized water to prepare an aqueous phase, the organic phase is gradually dripped into the aqueous phase under magnetic stirring at 700-900rpm, the magnetic stirring is carried out for 15 minutes, then ultrasonic emulsification is carried out for 3-7 s and 5 s at the gap under the ice bath condition by adopting a probe type ultrasonic crusher for 3 minutes at 360 w, then the emulsion is stirred and solidified at 700-900rpm for 10-14 hours at room temperature, the solidified BIT-PLA nanospheres are centrifuged for 25-35 minutes at 8000-12000 rpm and 22-28 ℃ for 3 times, the deionized water is washed and lyophilized to obtain BIT-PLA nanospheres, the BIT-PLA nanospheres are dispersed into 20 mL of chitosan solution with the mass fraction of 0.1-0.5%, and are stirred magnetically at 800 rpm for 12 hours, and the deionized water is washed for 3 times, and the chitosan-modified benzisothiazolinone polylactic acid nanospheres are prepared.
The second purpose of the invention is to provide the application of the chitosan modified benzisothiazolinone polylactic acid nanospheres in the preparation of antibacterial drugs.
The antibacterial drug is an antibioticE. coliAndS. aureusthe medicament of (1).
According to the invention, a solvent volatilization method is adopted to prepare the PLA nanosphere loaded with the BIT antibacterial drug, and chitosan is modified on the surface of the BIT-PLA nanosphere to obtain the chitosan-modified benzisothiazolinone polylactic acid nanosphere. The prepared chitosan-modified benzisothiazolinone polylactic acid nanospheres can effectively improve the transmembrane transport rate and antibacterial activity of a nano drug-loaded system, thereby being beneficial to the efficient utilization of benzisothiazolinone in the antibacterial fields of mildew prevention, corrosion prevention and the like and overcoming the problems of insolubility, dispersibility and stability of benzisothiazolinone.
Drawings
FIG. 1 is a hydrated particle size (A) of BIT-PLA nanospheres based on CS modification; (B) BIT-PLA-0.1% CS; (C) BIT-PLA-0.5% CS.
FIG. 2 is a zeta potential (A) BIT-PLA nanosphere based on CS modification of BIT-PLA; (B) BIT-PLA-0.1% CS; (C) BIT-PLA-0.5% CS.
FIG. 3 shows the fluorescent spectrum of (A) quantitative analysis of transmembrane transport fluorescence of coumarin-6 BIT-PLA-CS nanoparticle and (B) fluorescence value.
The specific implementation mode is as follows:
the following examples are further illustrative of the present invention and are not intended to be limiting thereof.
Example 1
(1) Dissolving 80 mg of polylactic acid PLA (average molecular weight 100 KDa) and 20 mg of benzisothiazolinone BIT raw drug in 5 mL of dichloromethane DCM to form an organic phase;
(2) Dissolving 120 mg of polyvinyl alcohol PVA (viscosity is 3.2-3.6, alcoholysis degree is 87.0% -89.0%) in 20 mL of deionized water to form a water phase;
(3) Slowly dripping the organic phase into the water phase under the magnetic stirring of 800 rpm, and stirring for 15 minutes by magnetic force;
(4) Ultrasonically emulsifying for 3 minutes by using a probe type ultrasonicator (JY 92-IIDN) under the ice bath condition of 360 w for 5 seconds and a gap of 5 seconds to form emulsion;
(5) Stirring and solidifying the emulsion at the room temperature at 800 rpm for 12 hours;
(6) Centrifuging the solidified BIT-PLA nanospheres for 30 minutes at the temperature of 10000 rpm and 25 ℃, washing the solidified BIT-PLA nanospheres for 3 times by using deionized water, and freeze-drying the solidified BIT-PLA nanospheres for later use to obtain the BIT-PLA nanospheres.
(7) And (2) adding 1 mg of coumarin 6 into the organic phase in the step (1), and repeating the steps 2-6 to prepare the fluorescent dye loaded coumarin 6 BIT-PLA.
The hydrated average particle size of the prepared BIT-PLA nanospheres was about 172.73 nm (FIG. 1A), corresponding to a zeta potential of-21.92 mV (FIG. 2A).
Example 2
(1) Dissolving 80 mg PLA (average molecular weight 100 KDa) and 20 mg BIT technical in 5 mL DCM to form an organic phase;
(2) Dissolving 120 mg of PVA (viscosity 3.2-3.6, alcoholysis degree 87.0% -89.0%) in 20 mL of deionized water to form a water phase;
(3) Slowly dripping the organic phase into the water phase under the magnetic stirring of 800 rpm, and stirring for 15 minutes by magnetic force;
(4) Ultrasonically emulsifying for 3 minutes by using a probe type ultrasonicator (JY 92-IIDN) under the ice bath condition of 360 w for 5 seconds and a gap of 5 seconds to form emulsion;
(5) Stirring and solidifying the emulsion at the room temperature at 800 rpm for 12 hours;
(6) Centrifuging the solidified BIT-PLA nanospheres at 10000 rpm and 25 ℃ for 30 minutes, and washing the solidified BIT-PLA nanospheres for 3 times by using deionized water;
(7) Weighing 20 mg of chitosan CS, and dissolving in 20 mL of deionized water solution (acetic acid, pH 5.0) to obtain CS solution (0.1%, w/v);
(8) Dispersing the BIT-PLA nanospheres in the step (6) into 20 mL of CS solution (0.1%, w/v, pH 5.0), stirring for 12 hours by magnetic force at 800 rpm, washing for 3 times by deionized water, preparing BIT-PLA-0.1% CS nanospheres (namely the chitosan-modified benzisothiazolinone polylactic acid nanospheres), and freeze-drying for later use.
(9) Adding 1 mg coumarin 6 to the organic phase of step (1), repeating steps 2-8, and making fluorescent dye-loaded coumarin 6 BIT-PLA-0.1% CS.
0.1% of the prepared BIT-PLA-0.1% of the hydrated average particle size of the CS nanospheres about 186.47 nm (FIG. 1B), corresponding to a zeta potential of 0.76 mV (FIG. 2B).
Example 3
(1) Dissolving 80 mg of PLA (average molecular weight of 100 KDa) and 20 mg of BIT technical in 5 mL of DCM to form an organic phase;
(2) Dissolving 120 mg of PVA (viscosity 3.2-3.6, alcoholysis degree 87.0% -89.0%) in 20 mL of deionized water to form a water phase;
(3) Slowly dripping the organic phase into the water phase under the magnetic stirring of 800 rpm, and stirring for 15 minutes by magnetic force;
(4) Ultrasonically emulsifying for 3 minutes by using a probe type ultrasonicator (JY 92-IIDN) under the ice bath condition of 360 w for 5 seconds and a gap of 5 seconds to form emulsion;
(5) Stirring and solidifying the emulsion at the room temperature at 800 rpm for 12 hours;
(6) Centrifuging the solidified BIT-PLA nanospheres at 10000 rpm and 25 ℃ for 30 minutes, and washing the solidified BIT-PLA nanospheres for 3 times by using deionized water;
(7) 100 mg of CS was weighed and dissolved in 20 mL of deionized water solution (acetic acid, pH 5.0) to give a CS solution (0.5%, w/v);
(8) Dispersing the BIT-PLA nanospheres in the step (6) into 20 mL of CS solution (0.5%, w/v, pH 5.0), stirring for 12 hours by magnetic force at 800 rpm, washing for 3 times by deionized water, preparing BIT-PLA-0.5% CS nanospheres (namely the chitosan-modified benzisothiazolinone polylactic acid nanospheres), and freeze-drying for later use.
(9) Adding 1 mg of coumarin 6 into the organic phase in the step (1), repeating the steps 2-8, and preparing the fluorescent dye-loaded coumarin 6 BIT-PLA-0.5 percent of CS.
0.5% of the prepared BIT-PLA-0.5% of the hydrated average particle size of the CS nanospheres of about 206.57 nm (FIG. 1C), corresponding to a zeta potential of 25.61 mV (FIG. 2C).
Example 4
Method for determining chitosan modified BIT-PLA nanospheres by adopting coumarin 6 fluorescent labeling methodE. coliAndS. aureustransmembrane transport capacity. 500. Mu.L of 2X 10 8 CFU/mL E. ColiOrS. aureusAddition to a 2 mL sterile centrifuge tube, followed by incubation with 500. Mu.L coumarin 6 fluorescently labeled BIT-PLA, BIT-PLA-0.1 CS or BIT-PLA-0.5 CS, with a final concentration of the BIT active ingredient of 2. Mu.g/mL, shaking at 37 ℃ and 180 rpm for 2 hours, followed by 3 washes with sterile phosphate buffered saline (PBS, 0.01M, pH 7.4), suspending the bacterial cells in 1 mL PBS, measuring by a fluorescence spectrometer (LS 45, perkin Elmer, MA, USA) at an excitation wavelength of 430 nm, with an emission wavelength scan ranging from 450 nm to 700 nmE. coliAndS. aureusfluorescence value of coumarin 6 in the cells (FIG. 3). Loading coumarin 6 BIT-PLA-0.1% CS and BIT-PLA-0.5% CS entryE. coliThe fluorescence values in the cells were about 1.08 times and 1.37 times of those in the BIT-PLA treated group, respectively. In contrast, load coumarin 6 BIT-PLA-0.1% CS and BIT-PLA-0.5%CS entryS. aureusThe in-vivo fluorescence values were about 1.15 and 1.43 times those of the BIT-PLA treated group, respectively. The positive charge CS interface modifies BIT-PLA nano-particles, so that the BIT-PLA nano-particles are easier to interact with negative charges on the surface of cells, and further, better permeability and transmembrane transport capacity are obtained.
Method for modifying BIT-PLA nanoparticles by using CS through turbidimetryE. coliAndS. aureusand (4) analyzing antibacterial activity. 100 μ L of concentration gradient BIT-PLA, BIT-PLA-0.1% CS or BIT-PLA-0.5% CS was added to the 96-well plate. Subsequently, 100. Mu.L of 2X 10 were inoculated per well 6 CFU/mL of the cells were cultured at 37 ℃ for 24 hours, and OD was measured by a microplate reader (Thermo Multiskan GO, thermo Fisher Scientific, varta, finland) 60 Calculate BIT-PLA, BIT-PLA-0.1% CS or BIT-PLA-0.5% CS pairs using IBM SPSS 25E. coliAndS. aureusEC of (1) 50 (tables 1 and 2). BIT-PLA, BIT-PLA-0.1% CS and BIT-PLA-0.5%E. coliAndS. aureus EC 50 the method comprises the following steps: 5.6331 μ g/mL,3.8305 μ g/mL and 1.9087 μ g/mL as well as 12.4093 μ g/mL,7.6955 μ g/mL and 5.8852 μ g/mL. Using as standard pharmaceutical agent BIT-PLA nanospheres without CS modification, BIT-PLA-0.5%E. coliAndS. aureusthe relative virulence is approximately 2.95 and 2.11 times that of BIT-PLA. The CS modified BIT-PLA nanospheres can effectively improve the antibacterial activity of the nano drug-loaded system.
Claims (4)
1. A preparation method of chitosan modified benzisothiazolinone polylactic acid nanospheres is characterized in that 70-90mg of polylactic acid (PLA) with an average molecular weight of 100 KDa and 20 mg of Benzisothiazolinone (BIT) are dissolved in 5 mL of dichloromethane to prepare an organic phase, 120 mg of polyvinyl alcohol is dissolved in 18-22 mL of deionized water to prepare an aqueous phase, the organic phase is gradually dripped into the aqueous phase under the magnetic stirring of 700-900rpm, the magnetic stirring is carried out for 15 minutes, then a probe type ultrasonicator is adopted to carry out ultrasonic emulsification for 3 minutes under the conditions of 360 w and ice bath for 3-7 seconds and 5 seconds of gap, ultrasonic emulsification is carried out for 3 minutes, then emulsion is stirred and solidified for 10-14 hours at 700-900rpm at room temperature, the solidified BIT-PLA nanospheres are placed in 8000-12000 rpm,22-28 ℃, the centrifugation is carried out for 25-35 minutes, the deionized water is washed for 3 times, freeze-drying is carried out to obtain the BIT-PLA nanospheres, the BIT-PLA nanospheres are dispersed into 20 mL of a chitosan solution with the mass fraction of 0.1-0.5%, the chitosan solution is modified by the magnetic stirring for 12 hours, and the chitosan nanospheres are washed by the deionized water to prepare the benzisothiazolinone polylactic acid nanospheres, and the chitosan nanospheres are washed by the deionized water, and the chitosan nanospheres are washed by stirring for 3 hours;
the viscosity of the polyvinyl alcohol is 3.2-3.6, and the alcoholysis degree is 87.0% -89.0%;
the chitosan solution is prepared by weighing chitosan and dissolving the chitosan in deionized water solution with acetic acid for adjusting the pH value to 5.0 to obtain chitosan solution;
the average particle size of the chitosan modified benzisothiazolinone polylactic acid nanosphere is 186.47-206.57 nm, and the corresponding zeta potential is 0.76-25.61 mV.
2. A chitosan-modified benzisothiazolinone polylactic acid nanosphere prepared according to the preparation method of claim 1.
3. The use of chitosan-modified benzisothiazolinone polylactic acid nanospheres as claimed in claim 2 in the preparation of antibacterial drugs.
4. The use according to claim 3, wherein said antibacterial agent is an antibioticE. coliOrS. aureusThe medicament of (1).
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN202110619426.5A CN113440523B (en) | 2021-06-03 | 2021-06-03 | Chitosan-modified benzisothiazolinone polylactic acid nanosphere and preparation method and application thereof |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN202110619426.5A CN113440523B (en) | 2021-06-03 | 2021-06-03 | Chitosan-modified benzisothiazolinone polylactic acid nanosphere and preparation method and application thereof |
Publications (2)
| Publication Number | Publication Date |
|---|---|
| CN113440523A CN113440523A (en) | 2021-09-28 |
| CN113440523B true CN113440523B (en) | 2022-12-30 |
Family
ID=77810602
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN202110619426.5A Active CN113440523B (en) | 2021-06-03 | 2021-06-03 | Chitosan-modified benzisothiazolinone polylactic acid nanosphere and preparation method and application thereof |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN113440523B (en) |
Families Citing this family (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN114113591B (en) * | 2021-11-17 | 2022-06-21 | 广东省科学院微生物研究所(广东省微生物分析检测中心) | Enzyme linked immunosorbent assay method for sensitively detecting Escherichia coli O157H7 |
Citations (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| WO2001080835A1 (en) * | 2000-04-24 | 2001-11-01 | Tanabe Seiyaku Co., Ltd. | Process for producing microsphere |
| CN100998566A (en) * | 2006-12-30 | 2007-07-18 | 南京工业大学 | Nanometer microsphere of temozolomide polylactic acid and its preparation and preparing method therefor |
| CN101889985A (en) * | 2010-07-08 | 2010-11-24 | 东华大学 | Medicament-carrying nano microspheres and preparation method thereof |
| CN107049984A (en) * | 2017-03-14 | 2017-08-18 | 武汉理工大学 | A kind of preparation method for carrying taxol polylactic-co-glycolic acid microballoon |
Family Cites Families (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| WO2011011978A1 (en) * | 2009-07-31 | 2011-02-03 | 西安力邦医药科技有限责任公司 | Nanosphere or microsphere drug carrier, preparation method, composition and use thereof |
-
2021
- 2021-06-03 CN CN202110619426.5A patent/CN113440523B/en active Active
Patent Citations (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| WO2001080835A1 (en) * | 2000-04-24 | 2001-11-01 | Tanabe Seiyaku Co., Ltd. | Process for producing microsphere |
| CN100998566A (en) * | 2006-12-30 | 2007-07-18 | 南京工业大学 | Nanometer microsphere of temozolomide polylactic acid and its preparation and preparing method therefor |
| CN101889985A (en) * | 2010-07-08 | 2010-11-24 | 东华大学 | Medicament-carrying nano microspheres and preparation method thereof |
| CN107049984A (en) * | 2017-03-14 | 2017-08-18 | 武汉理工大学 | A kind of preparation method for carrying taxol polylactic-co-glycolic acid microballoon |
Also Published As
| Publication number | Publication date |
|---|---|
| CN113440523A (en) | 2021-09-28 |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| Labib | Overview on zein protein: A promising pharmaceutical excipient in drug delivery systems and tissue engineering | |
| Khalil et al. | Bacterial-derived polymer poly-γ-glutamic acid (γ-PGA)-based micro/nanoparticles as a delivery system for antimicrobials and other biomedical applications | |
| Bashir et al. | Chitosan nanoparticles: a versatile platform for biomedical applications | |
| Cruz et al. | Antimicrobial activity of a new synthetic peptide loaded in polylactic acid or poly (lactic-co-glycolic) acid nanoparticles against Pseudomonas aeruginosa, Escherichia coli O157: H7 and methicillin resistant Staphylococcus aureus (MRSA) | |
| Su et al. | Preparation and properties of pH-responsive, self-assembled colloidal nanoparticles from guanidine-containing polypeptide and chitosan for antibiotic delivery | |
| Ji et al. | Binary and tertiary complex based on short-chain glucan and proanthocyanidins for oral insulin delivery | |
| Zheng et al. | Biodegradable and redox-responsive chitosan/poly (L-aspartic acid) submicron capsules for transmucosal delivery of proteins and peptides | |
| CN110801431B (en) | Construction and application of core-shell type intelligent nano delivery system | |
| CN106905443B (en) | The preparation method and applications of guanidinated chitosan | |
| CN110496229B (en) | Nanoparticle-coated antibacterial peptide with slow release property and preparation method thereof | |
| CN112089704A (en) | Bionic nano-carrier and preparation method and application thereof | |
| CN110623918A (en) | Carboxymethyl chitosan/sodium alginate nano hydrogel and preparation method and application thereof | |
| Nalini et al. | In vitro cytocompatibility assessment and antibacterial effects of quercetin encapsulated alginate/chitosan nanoparticle | |
| Chen et al. | Encapsulation of Phloretin in a ternary Nanocomplex prepared with Phytoglycogen–Caseinate–pectin via electrostatic interactions and chemical cross-linking | |
| CN113440523B (en) | Chitosan-modified benzisothiazolinone polylactic acid nanosphere and preparation method and application thereof | |
| Zhang et al. | Emamectin benzoate nanogel suspension constructed from poly (vinyl alcohol)-valine derivatives and lignosulfonate enhanced insecticidal efficacy | |
| CN111870579A (en) | Tumor-targeted nano micelle, preparation method and application of nano micelle as drug carrier | |
| Skirtenko et al. | A facile one-pot sonochemical synthesis of surface-coated mannosyl protein microspheres for detection and killing of bacteria | |
| Wang et al. | One-pot quaternization of dual-responsive poly (vinyl alcohol) with AIEgens for pH-switchable imaging and killing of bacteria | |
| CN111481673A (en) | Preparation method of multi-active-component preparation | |
| Yao et al. | Surface-modified polylactic acid nanospheres with chitosan for antibacterial activity of 1, 2-benzisothiazolin-3-one | |
| Gupta et al. | Primaquine loaded chitosan nanoparticles for liver targeting | |
| CN106399291A (en) | Galactosyl grafted-modified alginate microspheres and applications thereof | |
| Hadizadeh et al. | Amoxicillin-loaded polymeric nanoparticles of less than 100 nm: design, preparation and antimicrobial activity against methicillin-resistant staphylococcus aureus | |
| Salama | The development of a novel ultrashort antimicrobial peptide nanoparticles with potent antimicrobial effect |
Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| PB01 | Publication | ||
| PB01 | Publication | ||
| SE01 | Entry into force of request for substantive examination | ||
| SE01 | Entry into force of request for substantive examination | ||
| GR01 | Patent grant | ||
| GR01 | Patent grant |