CN109758582A - The carrier micelle delivery system that a kind of polyoxyethylene sorbitan monoleate penetrating blood-brain barrier and its component are formed - Google Patents
The carrier micelle delivery system that a kind of polyoxyethylene sorbitan monoleate penetrating blood-brain barrier and its component are formed Download PDFInfo
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Abstract
The invention discloses the carrier micelle delivery systems that a kind of polyoxyethylene sorbitan monoleate for penetrating blood-brain barrier and its component are formed.Application of the claimed polyoxyethylene sorbitan monoleate in the product that preparation can penetrate blood-brain barrier and the product that can penetrate blood-brain barrier containing polyoxyethylene sorbitan monoleate.Present invention discover that the micella that polyoxyethylene sorbitan monoleate and its component are formed can form carrier micelle with certain drugs for needing to penetrate blood-brain barrier but being difficult to penetrate blood-brain barrier, it is helped to penetrate blood-brain barrier, improve intracerebral drug concentration and bioavilability, to achieve the purpose that treat brain related disease, certain diseases at present still without medicine treatment are made to have the possibility for the treatment of, new Research Thinking and solution are provided for the research and difficult point of brain related disease, is of great significance to the exploration and development in the region.
Description
Technical field
The invention belongs to biological medical polymer material technical fields, and in particular to a kind of poly- sorb for penetrating blood-brain barrier
The carrier micelle delivery system that ester 80 and its component are formed.
Background technique
With the quickening increasingly of world population ages, the disease of nervous system is increasingly becoming the master for threatening human health
Problem is wanted, meanwhile, brain diseases also seriously endanger controlling for some brain diseases such as human health, especially brain tumor, cerebral ischemia
Treat one of the huge challenge in always medical development process.Its reason first is that many drugs cannot be by blood-brain barrier or logical
It excessively is not enough to control disease, increases Dose Toxicity and be incremented by.Blood-brain barrier is the weight for maintaining ambient stable in central nervous system
Institutional framework is wanted, effectively harmful substance can be prevented to enter in brain tissue, but will limit the transhipment of drug simultaneously, makes about 95%
Drug cannot enter brain from blood, be unfavorable for the treatment of disease.It is current for thus how increasing the bioavilability of drug
The major issue that nervous system disease faces.Current is some research shows that can be by changing medicines structure, changing
Become administration route, promote the opening of blood-brain barrier and increase the methods of the transhipment of drug to promote drug to pass through blood-brain barrier,
Prompt us that can take corresponding method according to the heterogeneity of drug, to reach the mesh of safe and effective treatment disease
's.
Currently, common pharmaceutical carrier organic and/or inorganic materials nanoparticle, dendritic, liposome, polymer nanocomposite
Hydrogel and polymer micelle etc..Wherein, polymer micelle is by the amphipathic copolymerization with hydrophilic segment and hydrophobic segment
Spontaneous assembling is formed object in water, has the advantage that polymer micelle has apparent nucleocapsid structure, in self assembly
Cheng Zhong, hydrophobic block constitutes the kernel of micella, and hydrophilic block then constitutes the shell of micella around micelle inner core, it will be apparent that
Nucleocapsid structure can effectively improve the water solubility of drug；Polymer latex beam stability is high, and partial size is small, uniform, has and overcomes blood
The function of brain barrier can pass through the methods of chemistry, physics and electrostatic interaction packet according to the heterogeneity of polymer hydrophobic segment
Hydrophobicity or poorly water soluble drugs are wrapped up in, hence into blood-brain barrier, improve drug concentration, stability and bioavilability, poison is secondary
It acts on small.Therefore blood-brain barrier is penetrated using polymer micelle as pharmaceutical carrier to have a good application prospect.
Many drugs cannot or seldom penetrate blood-brain barrier because of hydrophily or biggish relative molecular weight, clinically
Central nervous system disease cannot effectively be treated.Pharmaceutic adjuvant polyoxyethylene sorbitan monoleate is nonionic surface active agent, often in medicament
It is used as solubilizer in, the solubility of slightly solubility or fat-soluble medicine can be increased, can defend the oxidation and hydrolysis of drug, and to medicine
The stability of object preparation has protective effect.This of polyoxyethylene sorbitan monoleate and its component characteristic have biological therapy field potential
Auxiliary therapeutic action.
Summary of the invention
The object of the present invention is to provide the carrier micelles that a kind of polyoxyethylene sorbitan monoleate for penetrating blood-brain barrier and its component are formed
Delivery system.
Claimed polyoxyethylene sorbitan monoleate preparation can penetrate the application in the product of blood-brain barrier；
In addition, the product that can penetrate blood-brain barrier containing polyoxyethylene sorbitan monoleate shown in Formulas I, also belongs to protection of the invention
Range.
Specifically, the product concretely drug or pharmaceutical preparation or pharmaceutical carrier.
The drug concretely central nervous system depressants, central nervous system stimulant or brain tumor anticancer
Medicine；
The central nervous system depressants are more specifically selected from hypnotic sedative agent, Psychotolytic, antiepileptic, resist and shy
Any one fainted in medicine, antidepressants, anti-neurodegenerative disease medicine and antalgesic；
The central nervous system stimulant is chosen in particular from cerebral cortex stimulant, medullary respiratory center stimulant and promotion
Any one in brain function restorative；
Any one of the brain tumor anticarcinogen in chemotherapeutics and biological agent.
More specifically, the drug can be donepezil or Nimodipine.
The polyoxyethylene sorbitan monoleate concretely polyoxyethylene sorbitan monoleate micella, the poly- sorb containing polyoxyethylene fatty acid ester micella
Ester 80 contains the polysorbate by polyethylene glycol, the compound that polyoxyethylene alcohol acid anhydride and polyoxyethylene alcohol form or mixture
80。
A kind of pharmaceutical carrier or drug is also claimed in the present invention, and the pharmaceutical carrier or drug include polyoxyethylene sorbitan monoleate glue
Beam, polyoxyethylene fatty acid ester micella and the compound being made of polyethylene glycol, polyoxyethylene alcohol acid anhydride and polyoxyethylene alcohol mix
Close at least one of object.
Specifically, the pharmaceutical carrier or drug can for polyoxyethylene sorbitan monoleate micella, polyoxyethylene fatty acid ester micella and by
The compound or mixture of polyethylene glycol, polyoxyethylene alcohol acid anhydride and polyoxyethylene alcohol composition.
The pharmaceutical carrier or drug are specially the pharmaceutical carrier or drug that can penetrate blood-brain barrier.
The mode of appearance of the polyoxyethylene sorbitan monoleate micella and polyoxyethylene fatty acid ester micella is near-spherical structure；
The polyoxyethylene fatty acid ester micella is specially polyoxyethylene oleate micella.
The polyoxyethylene oleate micella is specially polyoxyethylene alcohol oleate micella or polyoxyethylene alcohol acid anhydride oleate
Micella.
The polyoxyethylene alcohol oleate micella is specially polyoxyethylene alcohol monoleate micella；
The polyoxyethylene alcohol acid anhydride oleate micella is specially polyoxyethylene alcohol acid anhydride monoleate micella or polyoxyethylene alcohol
Acid anhydride dioleate micella.
The polyoxyethylene alcohol monoleate micella is specially PIM micella；The PIM represents polyoxyethylene isosorbide
Oleate, molecular formula C24H42O5(OC2H4)n, n 5-27；Its structural formula is as shown in formula III；
The polyoxyethylene alcohol acid anhydride monoleate micella is specially PSM micella；The PSM represents polyoxyethylene sorbitol acid anhydride
Monoleate, molecular formula C24H44O6(OC2H4)n, n 15-33；Its structural formula is as shown in Formula II；
The polyoxyethylene alcohol acid anhydride dioleate micella is specially PSD micella；The PSD represents polyoxyethylene sorbitol acid anhydride
Dioleate, molecular formula C42H76O7(OC2H4)n, n 11-35；Its structural formula is as shown in formula IV.
It can be made according to various conventional methods when above-mentioned micella is as pharmaceutical carrier or drug, it such as can be according to including following step
Rapid method is made:
1) by polyoxyethylene sorbitan monoleate, polyoxyethylene fatty acid ester or by polyethylene glycol, polyoxyethylene alcohol acid anhydride and polyoxyethylene alcohol
After the mixture of composition is dissolved with sodium chloride injection, the solution I that concentration is 10mg/ml is obtained；
2) appropriate amount of drug to be loaded is dissolved with ethyl alcohol, obtains solution II；
3) solution I, solution II, ethyl alcohol and sodium chloride injection are mixed, evaporating ethanol is to get the polysorbate
80 micellas, polyoxyethylene fatty acid ester micella or the compound being made of polyethylene glycol, polyoxyethylene alcohol acid anhydride and polyoxyethylene alcohol.
Specifically, the concentration of the solution I can be 10mg/ml；
In the step 3), the volume ratio of solution I, ethyl alcohol and sodium chloride injection is 0.5:0.2:0.5.
In the evaporation step, temperature is 35 DEG C.
The polyoxyethylene sorbitan monoleate micella, polyoxyethylene fatty acid ester micella or by polyethylene glycol, polyoxyethylene alcohol acid anhydride and poly-
The compound of ethylene oxide alcohol composition or the load concentration of mixture are 0.3mg/ml.
It is described poly- in the compound or mixture being made of polyethylene glycol, polyoxyethylene alcohol acid anhydride and polyoxyethylene alcohol
Ethylene oxide alcohol acid anhydride is PS；The polyoxyethylene alcohol is PI；
The PS represents polyoxyethylene sorbitol acid anhydride, molecular formula C6H12O5(OC2H4)n, n 19-30；
The PI represents polyoxyethylene isobide, molecular formula C6H10O4(OC2H4)n, n 11-25；
The molecular formula of the polyethylene glycol is H (OCH2CH2)nOH, n 5-18.
Described PSM, PIM, PSD and the compound being made of polyethylene glycol, polyoxyethylene alcohol acid anhydride and polyoxyethylene alcohol mix
Closing object can be made according to the method included the following steps:
The tetrahydrofuran solution of the polyoxyethylene sorbitan monoleate is subjected to HPLC detection；
The condition of the HPLC detection is as follows: chromatographic column used is Gemini-NXC18 chromatographic column；
Eluent is made of mobile phase A and Mobile phase B；Wherein, mobile phase A is the first alcohol and water composition that volume ratio is 95:5
Mixed liquor；Mobile phase B is tetrahydrofuran；
Type of elution is gradient elution, elution speed 25mL/min；
The gradient elution mode is as follows:
To 59 seconds the 14th minute ends from 0th minute, eluent is mobile phase A；
To the 15.09th minute end from 15th minute, eluent is made of mobile phase A and Mobile phase B；The mobile phase A with
The volume ratio 50:50 of Mobile phase B；
To the 16.99th minute end from 15.1st minute, eluent is Mobile phase B；
To the 17.09th minute end from 17th minute, eluent is Mobile phase B；
To 18.99 minutes ends from 17.1st minute, eluent is mobile phase A；
To the 19th minute end from 19th minute, eluent is mobile phase A；
The component that retention time is 1.10-1.40 minutes is collected, removes solvent to get described by polyethylene glycol, polyoxy second
The compound or mixture of enol acid anhydride and polyoxyethylene alcohol composition；
The component that retention time is 1.90-2.25min is collected, removes solvent to get PSM；
The component that retention time is 2.70-3.30min is collected, removes solvent to get PIM；
The component that retention time is 4.50-6.60min is collected, removes solvent to get PSD；
Specifically, the concentration of the tetrahydrofuran solution of the polyoxyethylene sorbitan monoleate is 500mg/ml；
The length of the chromatographic column is 150 millimeters, and internal diameter is 21.2 millimeters, and the partial size of C18 is 5 μm, and aperture is
Nebulizer gas pressure is 60.0ps；Drift tube temperature is 60 DEG C；
The method for removing solvent is various conventional methods, is such as evaporated.
In addition, the aforementioned pharmaceutical carrier or drug, which is also claimed, in the present invention as pharmaceutical carrier or is preparing drug
Application and polyoxyethylene sorbitan monoleate micella, polyoxyethylene fatty acid ester micella in carrier or by polyethylene glycol, polyoxyethylene alcohol acid anhydride
The application in pharmaceutical carrier or drug is being prepared with the compound or mixture of polyoxyethylene alcohol composition.
The polyoxyethylene fatty acid ester is specially polyoxyethylene oleate.
The polyoxyethylene oleate is specially polyoxyethylene alcohol oleate or polyoxyethylene alcohol acid anhydride oleate.
The polyoxyethylene alcohol oleate is specially polyoxyethylene alcohol monoleate；
The polyoxyethylene alcohol acid anhydride oleate is polyoxyethylene alcohol acid anhydride monoleate or polyoxyethylene alcohol acid anhydride dioleate.
The polyoxyethylene alcohol monoleate is specially PIM；The PIM represents polyoxyethylene isobide monoleate,
Molecular formula is C24H42O5(OC2H4)n, n 5-27；
The polyoxyethylene alcohol acid anhydride monoleate is specially PSM；The PSM represents polyoxyethylene sorbitol acid anhydride list oleic acid
Ester, molecular formula C24H44O6(OC2H4)n, n 15-33；
The polyoxyethylene alcohol acid anhydride dioleate is specially PSD；The PSD represents two oleic acid of polyoxyethylene sorbitol acid anhydride
Ester, molecular formula C42H76O7(OC2H4)n, n 11-35.
The product that blood-brain barrier can be penetrated is specially by polyethylene glycol, polyoxyethylene alcohol acid anhydride and polyoxyethylene alcohol group
At compound or mixture.
It is described poly- in the compound or mixture being made of polyethylene glycol, polyoxyethylene alcohol acid anhydride and polyoxyethylene alcohol
Ethylene oxide alcohol acid anhydride is PS；The polyoxyethylene alcohol is PI；
The PS represents polyoxyethylene sorbitol acid anhydride, molecular formula C6H12O5(OC2H4)n, n 19-30；
The PI represents polyoxyethylene isobide, molecular formula C6H10O4(OC2H4)n, n 11-25；
The molecular formula of the polyethylene glycol is H (OCH2CH2)nOH, n 5-18.
The polyoxyethylene sorbitan monoleate may include following component: PSM, PIM, PSD, PEG, PS and PI；
The PSM represents Tween-80, molecular formula C24H44O6(OC2H4)n, n 15-33；Its
Structural formula is as shown in Formula II above；
The PIM represents polyoxyethylene isobide monoleate, molecular formula C24H42O5(OC2H4)n, n 5-27；Its
Structural formula is as shown in formula III above；
The PSD represents polyoxyethylene sorbitol acid anhydride dioleate, molecular formula C42H76O7(OC2H4)n, n 11-35；Its
Structural formula is as shown in formula IV above.
The PEG represents polyethylene glycol, and molecular formula is H (OCH2CH2)nOH, n 5-18；
The PS represents polyoxyethylene sorbitol acid anhydride, molecular formula C6H12O5(OC2H4)n, n 19-30；
The PI represents polyoxyethylene isobide, molecular formula C6H10O4(OC2H4)n, n 11-25.
In the polyoxyethylene sorbitan monoleate, except said components may also include following component:
Polyethylene glycol monooleate shown in Formula V (Polyethylene glycol monooleate, PM), PM molecular formula is
C18H33O2(OC2H4)nH, n 3-21；
The dioleate of polyoxyethylene isobide shown in Formula IV (Polyoxyethylene isosorbide dioleate,
PID), PID molecular formula is C42H74O6(OC2H4)n, n 6-25；The PID being esterified completely due to there is no hydrophilic hydroxyl in structure,
Therefore insoluble in water；
Glycol dioleate shown in Formula VII (Polyethylene glycol dioleate, PD), PD molecular formula is
C36H66O3(OC2H4)OnH, n 4-18；Degree of esterification is high, poor solubility；
The trioleate of polyoxyethylene sorbitol acid anhydride shown in Formula VIII (Polyoxyethylene sorbitan
Trioleate, PSTri), PSTri molecular formula is C60H108O8(OC2H4)n, n 17-35；Degree of esterification is high, soluble,very slightly；
Four oleate of polyoxyethylene sorbitol acid anhydride shown in Formula IX (Polyoxyethylene sorbitan
Tetraoleate, PSTetra), PSTetra molecular formula is C78H140O9(OC2H4)n, n 28-35；The PSTetra being esterified completely
It is insoluble in water due to there is no hydrophilic hydroxyl in structure.
The polyoxyethylene sorbitan monoleate is polyoxyethylene sorbitan monoleate commonly used in the art.The polyoxyethylene sorbitan monoleate can be from various commercial sources
It is commercially available.Such as it is purchased from Nanjing WeiEr chemical engineering Co., Ltd, batch number 20151203-3.Different commercial sources disclose out
The polyoxyethylene sorbitan monoleate sold contains PEG, PS, PI, PM, PID, PD, PSTri, but the concrete content of every kind of component may slightly not
Together.Such as be purchased from Nanjing WeiEr chemical engineering Co., Ltd, batch number be 20151203-3 polyoxyethylene sorbitan monoleate in, above-mentioned each component
Content is as shown in table 1, and the content of PM and PD is limited to detection method limitation and unlisted since itself is as micro.
The content table of one pack system in table 1, the polyoxyethylene sorbitan monoleate sample as obtained by area normalization method calculating
The product concretely drug or pharmaceutical preparation.
The drug concretely central nervous system depressants, central nervous system stimulant or brain tumor anticancer
Medicine；
The central nervous system depressants are more specifically selected from hypnotic sedative agent, Psychotolytic, antiepileptic, resist and shy
Any one fainted in medicine, antidepressants, anti-neurodegenerative disease medicine and antalgesic；
The central nervous system stimulant is chosen in particular from cerebral cortex stimulant, medullary respiratory center stimulant and promotion
Any one in brain function restorative；
Any one of the brain tumor anticarcinogen in chemotherapeutics and biological agent.
More specifically, the drug can be donepezil or Nimodipine.
It further include auxiliary material in the product；
The auxiliary material is chosen in particular from any one in polysorbate 20, polysorbate 40 and polysorbate 60.
The carrier micelle is the carrier micelle being prepared with physical method or chemical method；Physical method used or change
Method is conventional method.
The administration mode of the product that blood-brain barrier can be penetrated or drug concretely nasal-cavity administration, percutaneous dosing,
Ophthalmic administration, oral administration or drug administration by injection.
The carrier micelle delivery system that the polyoxyethylene sorbitan monoleate provided by the invention for penetrating blood-brain barrier and its component are formed, can
Applied in following administration route, drug is promoted to pass through, such as: nasal-cavity administration, percutaneous dosing, ophthalmic administration, oral administration and note
Penetrate administration.Nasal-cavity administration can be improved the intracerebral concentration of some drugs, at the same also have bioavilability is high, absorb rapidly, make
The advantages that with facilitating；Polymer micelle percutaneous dosing application mainly in terms of topical skin treating, be alternatively arranged as many lifes
The carrier of object macromolecular or gene, it is transdermal to play a role into whole body；Polymer micelle can be improved the solubility of therapeutic agent
And the bioavilability in eye, mitigate adverse reaction and is able to achieve active targeting administration；Polymer micelle be applied to take orally to
Medicine system helps to reduce drug to the irritation of gastrointestinal tract, increases the drug concentration of absorption site, improves drug in gastrointestinal tract
In stability, and then improve drug bioavailability；Injection polymer micelle good biocompatibility is applied to drug administration by injection
In have higher safety.
Present invention discover that the micella that polyoxyethylene sorbitan monoleate and its component are formed can be needed with certain through blood-brain barrier but difficulty
To form carrier micelle through the drug of blood-brain barrier, it is helped to penetrate blood-brain barrier, improves intracerebral drug concentration and biology benefit
Expenditure makes certain diseases at present still without medicine treatment have the possibility for the treatment of to achieve the purpose that treat brain related disease, this
It invents to be difficult to the drug through blood-brain barrier and providing a kind of new approach, is provided for the research and difficult point of brain related disease
New Research Thinking and solution is of great significance to the exploration and development in the region.
Detailed description of the invention
Fig. 1 be 0.1ng/ml donepezil reference substance solution MRM map (380.30 > 91.00,380.30 > 243.10,
380.30>362.20)。
Fig. 2 be 1.0ng/ml donepezil reference substance solution MRM map (380.30 > 91.00,380.30 > 243.10,
380.30>362.20)。
Fig. 3 be 10.0ng/ml donepezil reference substance solution MRM map (380.30 > 91.00,380.30 > 243.10,
380.30>362.20)。
Fig. 4 be polyoxyethylene sorbitan monoleate micella-donepezil MRM map (380.30 > 91.00,380.30 > 243.10,
380.30>362.20)。
Fig. 5 be PEG/PS/PI micella-donepezil MRM map (380.30 > 91.00,380.30 > 243.10,
380.30>362.20)。
Fig. 6 be PSM micella-donepezil MRM map (380.30 > 91.00,380.30 > 243.10,380.30 >
362.20)。
Fig. 7 be PIM micella-donepezil MRM map (380.30 > 91.00,380.30 > 243.10,380.30 >
362.20)。
Fig. 8 be PSD micella-donepezil MRM map (380.30 > 91.00,380.30 > 243.10,380.30 >
362.20)。
Fig. 9 is polyoxyethylene sorbitan monoleate micella-Nimodipine MRM map (419.30 > 343.10,419.30 > 301.05).
Figure 10 is PEG/PS/PI compound-Nimodipine MRM map (419.30 > 343.10,419.30 > 301.05).
Figure 11 is PSM micella-Nimodipine MRM map (419.30 > 343.10,419.30 > 301.05).
Figure 12 is PIM micella-Nimodipine MRM map (419.30 > 343.10,419.30 > 301.05).
Figure 13 is PSD micella-Nimodipine MRM map (419.30 > 343.10,419.30 > 301.05).
Figure 14 is the average grain diameter column diagram that laser particle analyzer measures different blank micellas and carrier micelle.
Figure 15 is the Morphological Characterization figure of determination of transmission electron microscopy difference blank micella and carrier micelle.
Figure 16 is tail vein injection polyoxyethylene sorbitan monoleate micella-fluorescence DiR mouse living imaging figure.
Figure 17 is the chromatogram that HPLC prepares polyoxyethylene sorbitan monoleate secondary component.
Specific embodiment
The present invention is further elaborated combined with specific embodiments below, but the present invention is not limited to following embodiments.Institute
State method is conventional method unless otherwise instructed.The raw material can obtain unless otherwise instructed from public commercial source.
Polyoxyethylene sorbitan monoleate used in following embodiments is purchased from Nanjing WeiEr chemical engineering Co., Ltd, and batch number 20151203-3 is secondary
The chromatogram of component is as shown in figure 17.
Embodiment 1,
Fluorescent dye DiR iodide, English name DiR iodide [1,1 '-dioctadecyl-3,3,3 ', 3 '-
Tetramethylindotricarbocyanine iodide], molecular weight 1013.39, excitation wavelength 740nm, launch wavelength
780nm, fat-soluble fluorescence probe, for marking cell membrane and hydrophobic tissue, with structure shown in Formula X:
Donepezil is hexahydropyridine type oxide, and chemical name is 1- benzyl -4- [(5,6- dimetlioxyindan ketone -2-
Base) methyl] piperidines, molecular formula C24H29NO3, molecular weight 379.49, pKa value (Strongest Acidic) is 17.02,
PKa value (Strongest Basic) is 8.62, and polarity isDonepezil belongs to fat-soluble medicine, is a kind of reversible
Acetylcholine enzyme inhibitor, have Formula XI shown in structure:
The chemical structural formula of Nimodipine, taxol and chlorogenic acid is respectively as shown in Formula XII, Formula XIII, Formula XIV:
Above-mentioned Tween 80-DiR micella, Tween 80-donepezil micella and Tween 80-Nimodipine micella can be according to various
Conventional method is made.It can be such as made according to the method included the following steps:
A1 10mg donepezil and Nimodipine) are taken respectively, is dissolved with ethyl alcohol, and the donepezil drug storage of 1mg/ml is made
The nimodipine medicament stock solution of standby liquid and 1mg/ml.
B1 10mgDiR) is taken, is dissolved with ethyl alcohol, the DiR stock solution of 1mg/ml is made.
C1 Tween 80 and its appropriate secondary component) are taken, is dissolved with sodium chloride injection, it is the molten of 10mg/ml that concentration, which is made,
Liquid.
D1 0.3mlDiR stock solution) is taken, until 0.2ml ethyl alcohol, the Tween 80 and its secondary of 0.5ml is added in the flask of former bottom
The solution and 0.5ml sodium chloride injection of component remove ethyl alcohol, it is molten that DiR micella are made with Rotary Evaporators in 35 DEG C of outstanding steamings
Liquid, wherein 1ml solution contains 0.3mgDiR.
E1 0.3ml donepezil medicine storage liquid and nimodipine medicament stock solution) are taken, until in round-bottomed flask, respectively plus
Enter 0.2ml ethyl alcohol, the solution of the Tween 80 of 0.5ml and its secondary component and 0.5ml sodium chloride injection, with Rotary Evaporators in
35 DEG C of outstanding steamings, remove ethyl alcohol, donepezil drug micelles solution and nimodipine medicament micellar solution are made, wherein 1ml solution
Donepezil containing 0.3mg and 0.3mg Nimodipine respectively.
Fig. 1 be 0.1ng/ml donepezil reference substance solution MRM map (380.30 > 91.00,380.30 > 243.10,
380.30>362.20)。
Fig. 2 be 1.0ng/ml donepezil reference substance solution MRM map (380.30 > 91.00,380.30 > 243.10,
380.30>362.20)。
Fig. 3 be 10.0ng/ml donepezil reference substance solution MRM map (380.30 > 91.00,380.30 > 243.10,
380.30>362.20)。
Fig. 4 be polyoxyethylene sorbitan monoleate micella-donepezil MRM map (380.30 > 91.00,380.30 > 243.10,
380.30>362.20)。
Fig. 5 be PEG/PS/PI micella-donepezil MRM map (380.30 > 91.00,380.30 > 243.10,
380.30>362.20)。
Fig. 6 be PSM micella-donepezil MRM map (380.30 > 91.00,380.30 > 243.10,380.30 >
362.20)。
Fig. 7 be PIM micella-donepezil MRM map (380.30 > 91.00,380.30 > 243.10,380.30 >
362.20)。
Fig. 8 be PSD micella-donepezil MRM map (380.30 > 91.00,380.30 > 243.10,380.30 >
362.20)。
Fig. 9 is polyoxyethylene sorbitan monoleate micella-Nimodipine MRM map (419.30 > 343.10,419.30 > 301.05).
Figure 10 is PEG/PS/PI compound-Nimodipine MRM map (419.30 > 343.10,419.30 > 301.05).
Figure 11 is PSM micella-Nimodipine MRM map (419.30 > 343.10,419.30 > 301.05).
Figure 12 is PIM micella-Nimodipine MRM map (419.30 > 343.10,419.30 > 301.05).
Figure 13 is PSD micella-Nimodipine MRM map (419.30 > 343.10,419.30 > 301.05).
In the present invention, the Senile Mouse of blank micella and carrier micelle method particularly includes:
A1 above-mentioned blank micella and carrier micelle) are taken, adds appropriate sodium chloride injection to dilute, uses laser particle analyzer
Measure its partial size.
B1 mode of appearance characterization) is carried out to above-mentioned blank micella and carrier micelle using transmission electron microscope (TEM), is taken
The micellar solution of debita spissitudo is added dropwise on special copper mesh, is dyed with 4% phosphotungstic acid, after spontaneously drying, is seen using TEM
Examine form.
In the present invention, the administration mode of mouse tail vein injection, the preparation method of biological sample are taken specifically:
A1 mouse tail vein injection, injection dosage 3mg/kg) are taken, every group of each 4 mouse removes positive controls mouse
Outside, 10min after administration, dissection take Mice brain tissues, and physiological saline is cleaned, dries, set -20 DEG C and save backup.Positive controls
Mouse, 2h after administration, dissection take Mice brain tissues, and physiological saline is cleaned, dries, set and measure fluorescence intensity in living imaging instrument.
B1 brain tissue will be weighed before) testing, and will smash to pieces, is added appropriate physiological saline (1g:2ml), and be vortexed and mix.It will homogenate
It after liquid is centrifuged 15min under the conditions of 10000rpm, 4 DEG C, pipettes 200 μ L of supernatant and is placed in 1.5ml tubule, 600 μ L first are added
Alcohol is vortexed, and mixes.Centrifugation, 10000rpm, 4 DEG C, 15min.Supernatant is pipetted in 1.5ml tubule, N2Drying.300 μ L are added
Methanol dissolves sediment, is vortexed and mixes.Centrifugation, 10000rpm, 4 DEG C, 15min.Supernatant is taken to carry out UPLC-MS/MS points
Analysis.
UPLC-MS/MS method is preferred specifically:
Instrument: SHIMADZU LC-MS 8050
CE: being 380.3 > m/z of m/z 91.00,380.3 > m/ of m/z for the qualitative ionic reaction of donepezil
380.3 > m/z of z243.10, m/z 362.20；It is 419.00 > m/z of m/z for the qualitative ionic reaction of Nimodipine
343.10, m/z 419.00 > m/z 301.05.
Ion source: ESI, positive ion detection, multiple-reaction monitoring (MRM) scanning
Ion source temperature: 300 DEG C
Chromatographic column: 2.6 μ C18 of Kimetex(50×2.1mm)
Flow velocity: 0.3ml/min
Column temperature: 30 DEG C
Mobile phase: A:5mM ammonium acetate aqueous solution B: acetonitrile；
Table 2, eluent gradient
In the present invention, living imaging instrument method specifically:
A1 1 mouse, intraperitoneal injection of anesthesia drug, injection dosage 0.1-0.15ml/20g) are taken at random.After anesthesia, tail
It is injected intravenously blank DiR solution, injection dosage 3mg/kg.After administration, 0min, 5min, 10min, 20min, 30min,
50min, 60min observe fluorescence distribution and fluorescence intensity, and mouse is dissected after 60min is administered, and take out Mice brain tissues, for sky
White group.
B1 7 mouse, intraperitoneal injection of anesthesia drug) are taken.After anesthesia, mouse tail vein injection polysorbate is randomly selected
80DiR carrier micelle, one pack system DIR polypeptide drug-loaded micelle solution, injection dosage 3mg/kg.After administration, 0min, 5min, 10min,
20min, 30min, 50min, 60min, 90min, 120min observe (excitation wavelength: 740nm, transmitted wave in living imaging instrument
Long 790nm) and the DiR fluorescence intensities of each group Mice brain tissues is calculated, it is experimental group.
In the present invention, the blank micella as made from film dispersion method and load donepezil, Nimodipine, chlorogenic acid and purple
The polyoxyethylene sorbitan monoleate of China fir alcohol and its partial size (as shown in figure 14) of one pack system PEG/PS/PI, PSM, PIM and PSD micella, as a result show
Show in addition to PEG/PS/PI, the blank micella partial size of polyoxyethylene sorbitan monoleate and its one pack system is in 200nm or less.PEG/PS/PI partial size
About 1.1 μm, and itself is without amphipathic, cannot form micella, to speculate that PEG/PS/PI and drug molecule may be in solution bodies
Composite construction is formed in system, this structure can improve insoluble drug dissolubility and it can be promoted to enter brain group through blood-brain barrier
It knits.And in carrier micelle, the load taxol drug micellar particle size of polyoxyethylene sorbitan monoleate and its one pack system is distributed between 5 μm -30 μm,
Partial size is excessive, thus it is speculated that may be because paclitaxel carried medicine micella is unstable, agglomeration is serious.Its appearance shape is detected by Electronic Speculum
State (as shown in figure 15), the blank micella and carrier micelle of polyoxyethylene sorbitan monoleate and its one pack system are near-spherical knot as the result is shown
Structure.
In the present invention, DiR micella that polyoxyethylene sorbitan monoleate and its one pack system PEG/PS/PI, PSM, PIM and PSD are prepared
Can be distributed in vivo and brain can be reached, thus it is speculated that polyoxyethylene sorbitan monoleate and its one-component PEG/PS/PI, PSM, PIM and
PSD may have Brain targeting effect.
In the present invention, schemed by polyoxyethylene sorbitan monoleate micella-donepezil and polyoxyethylene sorbitan monoleate micella-Nimodipine MRM
Spectrum and polyoxyethylene sorbitan monoleate micella-fluorescence DiR mouse living imaging figure (as shown in figure 16), it can be deduced that polyoxyethylene sorbitan monoleate glue
Beam-donepezil, polyoxyethylene sorbitan monoleate micella-Nimodipine and polyoxyethylene sorbitan monoleate micella-fluorescence DiR enter the blood brain of mouse
Barrier, thus it is speculated that may be because donepezil, Nimodipine and DiR contain theheterocyclic nitrogen atom on chemical structural formula, molecular weight exists
Between 300~400, it is easy to be wrapped up by micella.
In the present invention, there are four types of the drugs of mouse mainline, is respectively: donepezil, Nimodipine, taxol and green
Ortho acid show that donepezil and Nimodipine are able to enter the blood brain screen of mouse from the point of view of obtained mouse living imaging figure
Barrier.
Other two kinds of drug taxols and chlorogenic acid can not penetrate blood-brain barrier, thus it is speculated that may chemical structural formula with them
Related with relative molecular weight, without theheterocyclic nitrogen atom, relative molecular weight is respectively 418.44,853.9 and for taxol and chlorogenic acid
354.31.The pKa value of chlorogenic acid is 3.91, the micella that the acidity of itself may destroy polyoxyethylene sorbitan monoleate and its one pack system is formed
Complete structure, cause enter brain tissue before occur chlorogenic acid leakage.The pKa value of taxol is 11.9, alkaline drug
And molecular weight is larger, structure is complicated, thus it is speculated that on the one hand probably due to polyoxyethylene sorbitan monoleate and its micella of one pack system formation are in purple
It is unstable in the alkaline environment that China fir alcohol itself is formed, the complete structure of micella is destroyed, micella reunion is caused to show serious, partial size
It is excessive, it is difficult to pass through blood-brain barrier of mice；Another aspect taxane molecule size is larger to be difficult to by polyoxyethylene sorbitan monoleate and its single group
Subpackage is wrapped up in, and drug is caused to be easy leakage；The detection of instrument may also be lower than because of accumulation of the taxol in Mice brain tissues
Limit.
Claims (17)
1. application of the polyoxyethylene sorbitan monoleate in the product that preparation can penetrate blood-brain barrier.
2. application according to claim 1, it is characterised in that: the product be drug, pharmaceutical carrier, pharmaceutical preparation, in
Pivot nervous system depressant, central nervous system stimulant or brain tumor anticarcinogen.
3. the product that can penetrate blood-brain barrier containing polyoxyethylene sorbitan monoleate.
4. product according to claim 3, it is characterised in that: the product be drug, pharmaceutical carrier, pharmaceutical preparation, in
Pivot nervous system depressant, central nervous system stimulant or brain tumor anticarcinogen.
5. application according to any one of claims 1-4 or product, it is characterised in that: the polyoxyethylene sorbitan monoleate is poly- sorb
80 micella of ester, the polyoxyethylene sorbitan monoleate containing polyoxyethylene fatty acid ester micella or containing by polyethylene glycol, polyoxyethylene alcohol acid anhydride and
The compound of polyoxyethylene alcohol composition or the polyoxyethylene sorbitan monoleate of mixture.
6. a kind of pharmaceutical carrier or drug, including polyoxyethylene sorbitan monoleate micella, polyoxyethylene fatty acid ester micella and by polyethylene glycol,
At least one of the compound or mixture of polyoxyethylene alcohol acid anhydride and polyoxyethylene alcohol composition.
7. pharmaceutical carrier or drug described in claim 6 as pharmaceutical carrier or are preparing answering in pharmaceutical carrier or drug
With.
8. polyoxyethylene sorbitan monoleate micella, polyoxyethylene fatty acid ester micella or by polyethylene glycol, polyoxyethylene alcohol acid anhydride and polyoxyethylene
The compound or mixture of alcohol composition are preparing the application in pharmaceutical carrier or drug.
9. application described in pharmaceutical carrier according to claim 6 or drug or claim 7 or 8, it is characterised in that: institute
Blood-brain barrier can be penetrated by stating pharmaceutical carrier or drug.
10. according to the pharmaceutical carrier or drug or application any in claim 6-9, it is characterised in that: the poly- sorb
The mode of appearance of 80 micella of ester and polyoxyethylene fatty acid ester micella is near-spherical structure.
11. according to the pharmaceutical carrier or drug or application any in claim 5-10, it is characterised in that: the polyoxy second
Alkene aliphatic ester micella is polyoxyethylene oleate micella, polyoxyethylene alcohol oleate micella, polyoxyethylene alcohol acid anhydride oleic acid ester gum
Beam, polyoxyethylene alcohol monoleate micella, polyoxyethylene alcohol acid anhydride monoleate micella or polyoxyethylene alcohol acid anhydride dioleate micella.
12. pharmaceutical carrier according to claim 11 or application, it is characterised in that: the polyoxyethylene alcohol monoleate glue
Beam is PIM micella；The PIM represents polyoxyethylene isobide monoleate, molecular formula C24H42O5(OC2H4)n, n 5-
27；
The polyoxyethylene alcohol acid anhydride monoleate micella is PSM micella；The PSM represents polyoxyethylene sorbitol acid anhydride list oleic acid
Ester, molecular formula C24H44O6(OC2H4)n, n 15-33；
The polyoxyethylene alcohol acid anhydride dioleate micella is PSD micella；The PSD represents two oleic acid of polyoxyethylene sorbitol acid anhydride
Ester, molecular formula C42H76O7(OC2H4)n, n 11-35.
13. according to the pharmaceutical carrier or application any in claim 5-12, it is characterised in that: it is described by polyethylene glycol,
In the compound or mixture of polyoxyethylene alcohol acid anhydride and polyoxyethylene alcohol composition, the polyoxyethylene alcohol acid anhydride is PS；The polyoxy
Vinyl alcohol is PI；
The PS represents polyoxyethylene sorbitol acid anhydride, molecular formula C6H12O5(OC2H4)n, n 19-30；
The PI represents polyoxyethylene isobide, molecular formula C6H10O4(OC2H4)n, n 11-25；
The molecular formula of the polyethylene glycol is H (OCH2CH2)nOH, n 5-18.
14. according to the pharmaceutical carrier any in claim 5-13, it is characterised in that: described by polyethylene glycol, polyoxy second
The compound or mixture of enol acid anhydride and polyoxyethylene alcohol composition are made according to the method included the following steps:
The tetrahydrofuran solution of the polyoxyethylene sorbitan monoleate is subjected to HPLC detection；
The condition of the HPLC detection is as follows: chromatographic column used is Gemini-NXC18 chromatographic column；
Eluent is made of mobile phase A and Mobile phase B；Wherein, mobile phase A is the mixed of the first alcohol and water composition that volume ratio is 95:5
Close liquid；Mobile phase B is tetrahydrofuran；
Type of elution is gradient elution, elution speed 25mL/min；
The gradient elution mode is as follows:
To 59 seconds the 14th minute ends from 0th minute, eluent is mobile phase A；
To the 15.09th minute end from 15th minute, eluent is made of mobile phase A and Mobile phase B；The mobile phase A and flowing
The volume ratio 50:50 of phase B；
To the 16.99th minute end from 15.1st minute, eluent is Mobile phase B；
To the 17.09th minute end from 17th minute, eluent is Mobile phase B；
To 18.99 minutes ends from 17.1st minute, eluent is mobile phase A；
To the 19th minute end from 19th minute, eluent is mobile phase A；
Collecting retention time is 1.10-1.40 minute components, removing solvent to get；
Specifically, the concentration of the tetrahydrofuran solution of the polyoxyethylene sorbitan monoleate is 500mg/ml；
The length of the chromatographic column is 150 millimeters, and internal diameter is 21.2 millimeters, and the partial size of C18 is 5 μm, and aperture is
Nebulizer gas pressure is 60.0ps；Drift tube temperature is 60 DEG C.
15. according to claim the 2-14 any application or product or pharmaceutical carrier, it is characterised in that: during the drug is
Pivot nervous system depressant, central nervous system stimulant or brain tumor anticarcinogen.
16. any application or product or pharmaceutical carrier in -15 according to claim 1, it is characterised in that: the product or
It further include auxiliary material in drug；
The auxiliary material is chosen in particular from any one in polysorbate 20, polysorbate 40 and polysorbate 60.
17. any application or product or pharmaceutical carrier in -16 according to claim 1, it is characterised in that: described to wear
The product of saturating blood-brain barrier or the administration mode of drug are nasal-cavity administration, percutaneous dosing, ophthalmic administration, are administered orally or are administered to
Medicine.
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| CN111973750A (en) * | 2020-08-17 | 2020-11-24 | 中国食品药品检定研究院 | Polysorbate 80 product for optimizing PSM/PIM-oleate in different proportions and preparation method thereof |
| CN116120589A (en) * | 2022-12-26 | 2023-05-16 | 广西医科大学附属口腔医院 | Carboxylated polyamidoamine-amorphous calcium phosphate micelle and preparation method and application thereof |
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| CN116120589A (en) * | 2022-12-26 | 2023-05-16 | 广西医科大学附属口腔医院 | Carboxylated polyamidoamine-amorphous calcium phosphate micelle and preparation method and application thereof |
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