CN104208709A - Brain-targeted water soluble drug carrier as well as preparation method and application thereof - Google Patents

Brain-targeted water soluble drug carrier as well as preparation method and application thereof Download PDF

Info

Publication number
CN104208709A
CN104208709A CN201410423851.7A CN201410423851A CN104208709A CN 104208709 A CN104208709 A CN 104208709A CN 201410423851 A CN201410423851 A CN 201410423851A CN 104208709 A CN104208709 A CN 104208709A
Authority
CN
China
Prior art keywords
water soluble
soluble drug
ldl
brain targeting
brain
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.)
Withdrawn
Application number
CN201410423851.7A
Other languages
Chinese (zh)
Inventor
吴正治
段丽红
Current Assignee (The listed assignees may be inaccurate. Google has not performed a legal analysis and makes no representation or warranty as to the accuracy of the list.)
Individual
Original Assignee
Individual
Priority date (The priority date 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 date listed.)
Filing date
Publication date
Application filed by Individual filed Critical Individual
Priority to CN201410423851.7A priority Critical patent/CN104208709A/en
Publication of CN104208709A publication Critical patent/CN104208709A/en
Withdrawn legal-status Critical Current

Links

Landscapes

  • Pharmaceuticals Containing Other Organic And Inorganic Compounds (AREA)

Abstract

The invention provides a brain-targeted water soluble drug carrier as well as a preparation method and application thereof. An LDL (low density lipoprotein) receptor of an endothelial cell of blood-brain barrier is selected as a transmission water soluble drug to treat a target spot of central nervous system diseases, and chitosan with very good biocompatibility is taken as a drug carrier material to research a low-toxicity efficient water soluble drug nano brain-targeted preparation. The brain-targeted water soluble drug carrier can be used for strengthening brain targeting of water soluble drugs, strengthening the curative effect, lowering the toxic and side effect, and improving the bioavailability.

Description

A kind of Brain targeting water soluble drug load and preparation method thereof and application
Technical field
The invention belongs to targeting vector technical field of pharmaceuticals, particularly relate to a kind of Brain targeting water soluble drug load and preparation method thereof and application.
Background technology
Along with the aggravation of aged tendency of population process, the number that the whole world suffers from central nervous system (central nervous system, CNS) disease grows with each passing day.The treatment of many nervous system disease, needs to make medicine penetrate into whole central nervous system.
Blood brain barrier (blood-brain barrier, BBB) be one between blood and brain and spinal cord, the dynamic interface of lower, the selective handling capacity of permeability.The existence of blood brain barrier forms available protecting to cerebral tissue; avoid the infringement of harmful substance (as toxin and virus etc.); but also hinder many medicines to enter brain lesion district, as anticarcinogen, antibiotic and macromolecular substances (peptide class) etc.Its existence makes the medicine of 98% enter cerebral tissue, is the key factor of restriction medicine for central nervous system development.The medicine for central nervous system adopted clinically is the micromolecule liposoluble substance that can diffuse through blood brain barrier mostly, and this kind of medicine can not meet clinical needs far away, and diagnosis, the treatment of a lot of disease need macromole, water-soluble substances.Traditional method weak effect this kind of macromolecular drug being imported brain, danger are large, therefore in recent years for becoming focus gradually by the research of blood brain barrier Advances in Brain Targeting Drug Delivery.
Current solution medicine effectively has through the method for blood brain barrier: prepare prodrug, open blood brain barrier tight structure or use carrier system as antibody, liposome, nanoparticle etc.Wherein, prodrug exists and changes the property of medicine, introduces the risk of toxicity, and opens blood brain barrier tight structure and likely introduce other harmful substances.Carrier system of comparing becomes the more satisfactory method through blood brain barrier.
In carrier system, low density lipoprotein, LDL LDL is regarded as the natural partner of liposome, has lipid core, can be applied in the research of pharmaceutical carrier.LDL must be application as targeting factor in fields such as diagnosing tumor (load developing agent), antitumor drug and genophores.
According to the method that the treatment factor and LDL complex are formed, be mainly divided into two types: 1, treat the factor and insert LDL phospholipid layer by hydrophobic chain, thus " grappling " at LDL on the surface (" rivet method "); 2, the lipoid of LDL is extracted again with fat-soluble medicine restructuring LDL (" recombination method ").Wherein, in " rivet method ", the macromole containing hydrophobic side chain exists " embedding " LDL equally may.The people such as Jin-Seok Kim have studied by the application of " Terplex " system being with the polylysine/LDL/DNA of stearyl side chain to be formed as DNA vector, mainly reach stable by hydrophobic polylysine, electric charge between LDL and plasmid DNA/hydrophobic force balance, can't precipitate when ratio of components is suitable between system.The transfection efficiency of this system to smooth muscle A7R5 cell has even exceeded Lipofectin quite (exceeding 2.2 times), finds that the endocytosis of " Terplex " mediates via ldl receptor in an experiment.Further experimental result shows, still can be used as DNA vector in different cell line (CCD-32Lu fibroblast) this system.It is noted, however, that the ratio between polymer and LDL is extremely important, otherwise polymer can be caused to cover LDL completely.
The cholesterol ester at " recombination method " Shi Jiang LDL center extracts, then medicine is inserted in the Lipid monolayer ghost of LDL.This " recombination method " drawback is that operation is loaded down with trivial details, and Protein yield is not high, and whole process may destroy the structure of LDL, and requires to carry out hydrophobically modified to hydrophilic drugs.
Above result of study display LDL is playing a role as the ester gp that carries on the effect mainly LDL surface of targeting factor, and how to protect and carry an ester gp ApoB-100 to make it fully be exposed to cell membrane receptors surface be also the key that LDL realizes targeting.
Summary of the invention
The object of the present invention is to provide the load of a kind of Brain targeting water soluble drug, be intended to the brain targeting of enhancing water soluble drug, heighten the effect of a treatment, reduce its toxic and side effects, and improve its bioavailability.
Another object of the present invention is to the preparation method that the load of above-mentioned Brain targeting water soluble drug is provided.
Another object of the present invention is to the application that the load of above-mentioned Brain targeting water soluble drug is provided.
Another object of the present invention is to provide the Brain targeting aqueous solubility pharmaceutical formulations prepared by the load of above-mentioned Brain targeting water soluble drug.
Another object of the present invention is the preparation method providing above-mentioned Brain targeting aqueous solubility pharmaceutical formulations.
The present invention is achieved in that the load of a kind of Brain targeting water soluble drug, comprises pharmaceutical carrier propine acylation chitosan PA-CS-CHO, nitrine probe N-(2-(2-nitrine ethyoxyl) ethyl) palmitamide NAEP and LDL; LDL surface-assembled nitrine probe NAEP, the alkynyl on PA-CS-CHO and N 3nitrine on-LDL reacts and forms the amphipathic molecule of LDL targeting end as hydrophobic side.
Preferably, the structural formula of described propine acylation chitosan PA-CS-CHO is as follows:
Wherein, the scope of x is 2 ~ 30; The scope of y ' is 2 ~ 20; The scope of z is 8 ~ 30.
Invention further provides the preparation method of above-mentioned Brain targeting water soluble drug load, comprise the following steps:
By N-(2-(2-nitrine ethyoxyl) ethyl) palmitamide NAEP and low density lipoprotein, LDL LDL mixing in broad dose altogether, obtaining by self-assembling reaction the LDL that surface has nitrine probe, being labeled as N 3-LDL; Wherein, the mol ratio of described N-(2-(2-nitrine ethyoxyl) ethyl) palmitamide NAEP, low density lipoprotein, LDL LDL is 1: (0.5 ~ 2);
React under mild conditions by click chemistry, the alkynyl on propine acylation chitosan PA-CS-CHO and N 3nitrine on-LDL reacts, and obtains the load of Brain targeting water soluble drug; Wherein, described propine acylation chitosan PA-CS-CHO and N 3the mol ratio of-LDL is 1: (0.1 ~ 2).
Preferably, the preparation of described N-(2-(2-nitrine ethyoxyl) ethyl) palmitamide NAEP comprises the following steps:
(1) C 1sh 31cONHCH 2cH 2oCH 2cH 2the synthesis of OH (NHEP)
In a nitrogen atmosphere, dimethyl formamide stirring and dissolving is added in Palmic acid, 2-(2-amino ethoxy) ethanol and triethylamine mixed liquor, after ice bath cooling, add the BTA-N of mixing in advance, N, N ', N '-tetramethylurea hexafluorophosphate and N-hydroxy benzo triazole, room temperature reaction, thin layer chromatography determination reaction end, obtains reactant solution; Wherein, described Palmic acid, 2-(2-amino ethoxy) ethanol, triethylamine, dimethyl formamide, BTA-N, N, the mol ratio of N ', N '-tetramethylurea hexafluorophosphate and N-hydroxy benzo triazole is 1: (1 ~ 1.5): (1 ~ 3): (1 ~ 5): (1 ~ 4.5): (1 ~ 6);
Reactant solution is dropwise instilled in the cold water in stirring, with chloroform extraction; The chloroformic solution collected uses citric acid solution successively, sodium bicarbonate solution, and saturated nacl aqueous solution washs, and revolve after anhydrous sodium sulfate drying and boil off chloroform and obtain crude product, crude product is recrystallization purifying in ethanol, vacuum drying, and exsiccator is preserved;
(2) C 15h 31cONHCH 2cH 2oCH 2cH 2the synthesis of OTs (PEMBS)
The NHEP that step (1) obtains is dissolved in chloroform, adds pyridine; After cooling, add p-methyl benzene sulfonic chloride, return back to room temperature, stirring is spent the night, and thin layer chromatography determination reaction end, obtains product; Wherein, the mol ratio of described NHEP, chloroform, pyridine, p-methyl benzene sulfonic chloride is 1: (1 ~ 6): (4 ~ 5.6): (1 ~ 1.4);
Product instilled in hydrochloric acid solution, centrifugal going out after supernatant obtains product; Product rinses with hydrochloric acid and water respectively, at room temperature vacuum drying, and exsiccator stores;
(3) C 15h 31cONHCH 2cH 2oCH 2cH 2n 3(NAEP) synthesis
The PEMBS obtained in step (2) is dissolved in dimethyl formamide under nitrogen protection, adds sodium azide, slowly heat, after backflow certain hour, be down to room temperature; Reaction mixture instills in frozen water in stirring, filters, and after water repeatedly flush cake, room temperature in vacuo is dry; Obtain end product by column chromatography purification, at room temperature vacuum drying, exsiccator stores; Wherein, the mol ratio of described PEMBS, dimethyl formamide, sodium azide is 1: (1 ~ 5): (4 ~ 5.6).
Preferably, the preparation of described propine acylation chitosan PA-CS-CHO comprises the following steps:
Under nitrogen protection, the NaAc_HAc buffer solution of chitosan and Potassium metaperiodate. are with stirring reaction at suitable ratio 4 DEG C; Then in solution, add appropriate ethylene glycol solution carry out stopped reaction; Product dialyses to remove impurity respectively after rotary evaporation is concentrated in NaCl aqueous solution and deionized water, and end product lyophilization, obtains hydroformylation chitosan CS-CHO; Wherein, the molal volume ratio between described chitosan, NaAc_HAc buffer solution, Potassium metaperiodate., ethylene glycol is 1: (1 ~ 5): (0.1 ~ 1): (0.1 ~ 1);
Under nitrogen protection, in acetylenecarboxylic acid aqueous solution, add N-hydroxy-succinamide, dropwise instill in hydroformylation chitosan CS-CHO aqueous solution after dissolving, after stirring, add 1-ethyl-(3-dimethylaminopropyl) carbodiimide hydrochloride; The mol ratio of hydroformylation chitosan, acetylenecarboxylic acid, N-hydroxy-succinamide and 1-ethyl-(3-dimethylaminopropyl) carbodiimide hydrochloride is 1: (1 ~ 4): (1 ~ 3): after (1 ~ 3) mixture reaction 10 ~ 72h, be transferred in bag filter, the unreacted micromolecule of dialysis removing in NaCl aqueous solution and deionized water, lyophilization obtains product.
Invention further provides a kind of Brain targeting aqueous solubility pharmaceutical formulations, comprise the load of above-mentioned Brain targeting water soluble drug and Brain targeting water soluble drug; Wherein, the exposed Brain targeting aqueous solubility pharmaceutical formulations outside of LDL targeting end is formed as the amphipathic molecule of hydrophobic side and water soluble drug self assembly in the load of Brain targeting water soluble drug.
Preferably, the form of described Brain targeting aqueous solubility pharmaceutical formulations is the Nano microsphere of 35 ~ 100 nanometers; Described Brain targeting water soluble drug is the diagnosis of central nervous system disease, macromole, the water-soluble substances for the treatment of needs, comprises lycoramine, galantamine, lycorine.
Invention further provides the preparation method of above-mentioned Brain targeting aqueous solubility pharmaceutical formulations, comprise the following steps: the load of Brain targeting water soluble drug is dissolved in organic solvent and prepares organic solution, then the aqueous solution of described organic solution and Brain targeting water soluble drug is carried out self assembly in bag filter; Wherein, the volume of the load of Brain targeting water soluble drug and organic solvent is 1: (1 ~ 10), and the WMCO=1000 of bag filter, described organic solvent comprises dimethyl sulfoxide, DMF, acetone, oxolane.
Invention further provides the load of above-mentioned Brain targeting water soluble drug and prepare the application in Brain targeting aqueous solubility pharmaceutical formulations.
Invention further provides the load of above-mentioned Brain targeting water soluble drug and prepare the application in central nervous system disease medicine.
The present invention overcomes the deficiencies in the prior art, a kind of Brain targeting water soluble drug load and preparation method thereof and application are provided, treated the target spot of central nervous system disease as transmission water soluble drug by the ldl receptor of choosing blood-brain barrier endothelial cell, using the good chitosan of biocompatibility as drug carrier material, development low toxicity efficient water soluble drug nanometer Brain targeting preparation, its objective is the brain targeting of enhancing water soluble drug, heighten the effect of a treatment, reduce its toxic and side effects, and improve its bioavailability.
Accompanying drawing explanation
Fig. 1 be CS in the embodiment of the present invention 1 (on) and CS-CHO (under) 1h NMR spectrogram;
Fig. 2 be CS in the embodiment of the present invention 1 (under) and CS-CHO (on) FT-IR spectrogram;
Fig. 3 is the UV spectrogram of CS-CHO (dotted line) and PA-CS-CHO (solid line) in the embodiment of the present invention 1;
Fig. 4 be CS-CHO in the embodiment of the present invention 1 (on) and PA-CS-CHO (under) FT-IR spectrogram;
Fig. 5 is PA-CS-CHO and CS-CHO in the embodiment of the present invention 1 1h NMR spectrogram;
Fig. 6 is Palmic acid (a), C in the embodiment of the present invention 2 15h 31cONHCH 2cH 2oCH 2cH 2oH (b), C 15h 31cONHCH 2cH 2oCH 2cH 2oTs's (c) and NAEP (d) 1h NMR spectrogram;
Fig. 7 is the synthesis schematic diagram of Brain targeting water soluble drug load in the embodiment of the present invention 4.
Detailed description of the invention
In order to make object of the present invention, technical scheme and advantage clearly understand, below in conjunction with drawings and Examples, the present invention is further elaborated.Should be appreciated that specific embodiment described herein only in order to explain the present invention, be not intended to limit the present invention.
Embodiment 1
1, the synthesis of hydroformylation chitosan (CS-CHO)
As a kind of natural polysaccharide, the features such as chitosan has good biocompatibility, biodegradable, are particularly suitable for pharmaceutical carrier.Then the subject matter limiting its application is that water solublity is poor, needs could dissolve in acid medium.Therefore in this project, first utilize periodate to carry out oxidation processes to chitosan, obtain containing adjacent dialdehyde functional group on different molecular weight, chitosan chain, substantially increase its water solublity.Synthetic route is as follows:
Wherein, the scope of x is 2 ~ 30; The scope of y is the scope of 10 ~ 50, y ' is 2 ~ 20; The scope of z is 8 ~ 30, wherein y=y '+z.
Under nitrogen protection, in acetylenecarboxylic acid aqueous solution, add N-hydroxy-succinamide, dropwise instill in hydroformylation chitosan CS-CHO aqueous solution after dissolving, after stirring, add 1-ethyl-(3-dimethylaminopropyl) carbodiimide hydrochloride; Hydroformylation chitosan, acetylenecarboxylic acid, N-hydroxy-succinamide and 1-ethyl-(, 3-dimethylaminopropyl) mol ratio of carbodiimide hydrochloride is 1: (1 ~ 4): (1 ~ 3): after (1 ~ 3) mixture reaction 10 ~ 72h, be transferred in bag filter, the unreacted micromolecule of dialysis removing in NaCl aqueous solution and deionized water, lyophilization obtains product.
Take 29.84 grams of sodium acetate trihydrates and measure the acetic acid of 27.22 milliliter 36% in beaker, add the distilled water diluting of 200 ~ 300 milliliters, be stirred to and dissolve completely, be poured in the volumetric flask of 1 liter, use distilled water standardize solution again, solution is poured in washed reagent bottle, obtains the NaAc_HAc buffer solution of pH=4.5.
Under nitrogen atmosphere, 1mol chitosan is dissolved in the NaAc_HAc buffer solution of 1mol, obtains the NaAc_HAc buffer solution of chitosan, by the NaAc_HAc buffer solution of chitosan and 0.1mol Potassium metaperiodate. (KIO 4) with stirring reaction at suitable ratio 4 DEG C.Then the ethylene glycol solution adding 1mol in solution carrys out stopped reaction.Product dialyses to remove impurity respectively after rotary evaporation is concentrated in NaCl aqueous solution and deionized water.End product lyophilization, for subsequent use.
Oxidation product CS-CHO is characterized respectively with FI-IR and NMR.From the nuclear magnetic spectrogram of Fig. 1, H-3 in CS, the chemical shift of 4,5,6,6 ' appears at 3.3 ~ 3.8ppm, and H-1 appears at 4.6ppm, and H-2 is at about 2.8ppm place.The proton peak of aldehyde radical has been there is at 8.0ppm in the spectrogram of CS-CHO, and H-3, the integral area of 4,5,6,6 ' and the integral area ratio of H-2 are increased to 5.11 by 4.58 of CS.
Can find out that from infrared spectrum and Fig. 2 chitosan is at 1590cm -1there is amino bending vibration peak at wave number place, shows that chitosan has the order be not acylated by amino, 3426cm -1it is the absworption peak of hydroxyl.After Potassium metaperiodate. oxidation, there is the 1523cm of obvious feature -1and 1640cm -1peak, is respectively amide II peak and amide I peaks.And 2760 and 2780cm -1equal bimodal of intensity that place occurs and at 1660cm -1there is absworption peak at place, can judge that CS-CHO has aldehyde radical thus.
2, the synthesis of propine acylation chitosan (PA-CS-CHO)
After periodate oxidation, the water solublity of CS-CHO increases substantially, and can be dissolved in deionized water well, therefore can modify it in equal phase medium.Adopt NHS activated carboxyl, EDC hydrochlorate, as condensing agent, carries out the reaction of " treating different things alike " in aqueous medium.All reactants and product are all water-soluble, and small molecular weight impurity can remove in dialysis.Concrete synthetic route is as follows:
Wherein, the scope of x is 2 ~ 30; The scope of y ' is 2 ~ 20; The scope of z is 8 ~ 30.
Under nitrogen protection; in 4mol acetylenecarboxylic acid aqueous solution; add 1molN-N-Hydroxysuccinimide (NHS); dropwise instill after dissolving in 1mol hydroformylation chitosan CS-CHO aqueous solution, after stirring, add 3moll-ethyl-(3-dimethylaminopropyl) carbodiimide hydrochloride (EDC-HCL).After mixture reaction 10 ~ 72h, be transferred in bag filter, the unreacted micromolecule of dialysis removing in NaCl aqueous solution and deionized water.Lyophilization obtains product, for subsequent use.
Determine PA-CS-CHO and CS-CHO with UV spectrogram and there is different chemical constitutions; as shown in Figure 3; can see from spectrogram; after propine acylation reaction occurs; the maximum absorption wavelength of product moves to 275nm from 325nm; and the fine structure at 325 ~ 350nm place disappears; this is because after amino is acylated; its lone pair electrons and propioloyl form conjugated system; for sugared ring, the n that amino provides → pi-electron transition ability reduces, and makes wavelength blue shift; and for propine group, substituent group makes it absorb red shift.Its common result causes the change of uv absorption wavelength.
The FT-IR spectrogram of CS-CHO and PA-CS-CHO also also exists difference, as shown in Figure 4, can see, at 2200cm from spectrogram -1there is the stretching vibration peak of faint alkynyl in place's (see circled), proves the existence of alkynes.And in CS-CHO the C-N stretching vibration of free amino group from ~ 1500cm -1place's (see small arrow) disappears, and the substitute is ~ 1400cm -1the amide C-N stretching vibration at place absorbs.
In the present invention, utilize further 1h NMR demonstrates the structure of PA-CS-CHO, as shown in Figure 5, from 1h NMR spectrogram, after (be standard with methyl absworption peak on acetyl group (~ 1.9ppm, vertical line place)) propine acylation reaction occurs, the proton uptake peak of H-2 is moved to 2.85ppm from 3.0ppm.This is because after amino is acylated, the hydrogen bond formed with solvent reduces and weakens.Different alternately due to from solvent of the molecular weight of polymer, residing chemical fields is different, and the displacement therefore causing proton uptake peak is different.
Can compare with CS spectrogram (see Fig. 2), the CS H-2 absworption peak of 88% de-acetyl is near 2.85ppm, and after hydroformylation, due to molecular melting performance enhancement, the hydrogen bond effect between intermolecular and solvent strengthens, and H-2 absworption peak moves to 3.0ppm to low field.Again propine acidylate, H-2 has got back to 2.85ppm place.Although notice the excessive twice of acetylenecarboxylic acid, propine acylation reaction can not reach 100%, and this point also can to find out from splitting of H-2 peak.The absorption of alkynyl hydrogen H-a appears at ~ 2.75ppm place (see small arrow).
The synthesis of embodiment 2 nitrine probe
" probe " can be fixed in the phospholipid layer of LDL by the Palmic acid with cetyl, reactivity is lost in order to prevent functional group-nitrine end group from being imbedded in LDL completely, we introduce a single diglycol ethylene unit as " spacer ", synthesize N-(2-(2-nitrine ethyoxyl) ethyl) palmitamide (NAEP).Concrete synthetic route is as follows:
Wherein, a, 2-(2-amino ethoxy) ethanol, HBTU and HOBt of 1N, room temperature reaction 24 hours in DMF solution; B, TsCl, pyridine; C, 1.2N sodium nitride, pyridine.
(1) C 15h 31cONHCH 2cH 2oCH 2cH 2the synthesis of OH (NHEP)
By 1mol Palmic acid, 1mol 2-(2-amino ethoxy) ethanol and 3mol triethylamine in a nitrogen atmosphere, add 3mol dimethyl formamide (DMF) stirring and dissolving, insert in ice bath, 2mol BTA-the N of mixing is in advance added after cooling, N, N ', N '-tetramethylurea hexafluorophosphate (HBTU) and 3molN-hydroxy benzo triazole (HOBt), room temperature reaction.
Thin layer chromatography determination reaction end.Reactant solution dropwise instills in the cold water in stirring, uses chloroform extraction.The chloroformic solution collected uses citric acid solution successively, sodium bicarbonate solution, and saturated nacl aqueous solution washs, and revolves and boil off chloroform and obtain crude product after anhydrous sodium sulfate drying.Crude product is recrystallization purifying in ethanol, vacuum drying, and exsiccator is preserved.
(2) C 1sh 31cONHCH 2cH 2oCH 2cH 2the synthesis of OTs (PEMBS)
1mol NHEP is dissolved in 5mol chloroform, adds 5mol pyridine; After cooling, add 1.2mol p-methyl benzene sulfonic chloride, return back to room temperature, stirring is spent the night.Thin layer chromatography determination reaction end.In product instillation hydrochloric acid solution, centrifugal going out after supernatant obtains product.Product rinses with hydrochloric acid and water respectively, at room temperature vacuum drying, and exsiccator stores.
(3) C 15h 31cONHCH 2cH 2oCH 2cH 2n 3(NAEP) synthesis
1mol PEMBS is dissolved under nitrogen protection in 5mol dimethyl formamide (DMF), adds 5mol sodium azide (NaN 3), slowly heat, after backflow certain hour, be down to room temperature.Reaction mixture instills in frozen water in stirring, filters, and after water repeatedly flush cake, room temperature in vacuo is dry.Obtain end product by column chromatography purification, at room temperature vacuum drying, exsiccator stores.
1h NMR demonstrates completing (as shown in a, b in Fig. 6) of amidation process.After reacting, at 3.0-4.0ppm, (there is the characteristic absorption peak of tirethylene glycol unit four methene protons in e, f, g) place.Owing to being amidated, the electronegativity of carbonyl carbon reduces, and the methene proton absworption peak d of next-door neighbour's carboxyl moves to d ' to High-Field.The integration ratio of absworption peak is:
: c ': d ': (e+f+g)=1.54: 11.48: 1.08: 1.00: 4.25, coincidence theory value a ': b '.
From 1h NMR result (as fig. 6 c), after sulfonic acid esterification occurs, the characteristic absorption of phenyl ring has been there is between 7.0 ~ 8.0ppm, and the methene proton absworption peak adjacent with terminal hydroxy group moves to 4.1ppm (d ') from 3.9ppm (d), simultaneously, the absworption peak (2.4ppm, e) of former hydroxyl Labile protons disappears, and confirms the generation of esterification.
Weak impurity peaks residual between 3.0 ~ 4.0ppm shows sulfonic acid esterification and not exclusively, we are 80% by integral and calculating sulphonation rate.Because not esterified hydroxyl does not substantially react in next step reaction, therefore we do not carry out further purification to it.
1h NMR spectrogram (as shown in fig 6d) shows, after substitution reaction occurs, and characteristic absorption disappearance (7.0 ~ 8.0ppm:e, the f of tosylate groups; 2.4ppm, g).Give electronic effect due to azido group, the methene proton of next-door neighbour's end group absorbs and significantly moves to 3.85ppm (d ') from 4.1ppm.A, b, c tri-methine protons absworption peaks also move to High-Field to some extent.
The probe mark of embodiment 3 LDL
Adopt density gradient method (list of references: Lundberg, B.Preparation of drug-low density lipoprotein complexes for delivery of antitumoral drugs via the low density lipoprotein pathway.Cancer Res.1987,47,4105) from human plasma, LDL is extracted.NAEP and LDL in mixing in broad dose altogether, by self-assembling reaction obtain surface have nitrine probe LDL (NAEP-LDL, mark oneself be N 3-LDL).
Particle size distribution and zeta current potential between NAEP-LDL and primary LDL after compound, as shown in table 1 below:
The composition of original LDL and NAEP-LDL of table 1, mean diameter and zeta current potential
Can find according to table 1, after compound, between NAEP-LDL and primary LDL, particle size distribution and zeta current potential are more or less the same, and illustrate that hydrophobic probe is modified and do not change too much the mobility of LDL in water.This point is extremely important, because the change that LDL surface potential occurs can promote that scavenging cells cell is to its endocytosis, thus is removed rapidly from blood circulation by LDL.
The preparation of embodiment 4 water soluble drug nanometer Brain targeting preparation
The mol ratio of the load of Brain targeting water soluble drug and organic solvent is 1: (1 ~ 10), and the WMCO=1000 of bag filter, described organic solvent comprises dimethyl sulfoxide, DMF, acetone, oxolane.
Alkynyl on PA-CS-CHO under mild conditions and N is reacted by click chemistry 3nitrine on-LDL reacts and synthesizes the amphipathic molecule of LDL targeting end as hydrophobic side.Concrete synthetic route is as follows:
This molecule and the self assembly of water soluble drug maryllidaceous alkaloid form the exposed water soluble drug nanometer Brain targeting preparation outside of LDL targeting end, and as shown in Figure 7, after self assembly, the form of preparation is the Nano microsphere of 35 ~ 100 nanometers to assembling process.
The concrete steps of self assembly: the dimethyl sulfoxide DMSO solution pipetting 20 μ L amphipathic molecules with micropipettor instills in 2 μ L maryllidaceous alkaloid solution, after shaking up, 37 DEG C of vibration 12h.Be transferred in bag filter (WMCO=1000), in the PBS buffer containing 0.01%EDTA, dialyse at 4 DEG C 24h, exchange buffering liquid three times.After filtering filter membrane with 0.22 μm, 4 DEG C of preservations.
Compared to the shortcoming and defect of prior art, the present invention has following beneficial effect:
1, the method operation of water soluble drug and LDL compound that adopts of the present invention is simple, to make a year ester gp ApoB-100 fully be exposed to cell membrane receptors surface.
2, to develop the toxic and side effects of water soluble drug nanometer Brain targeting preparation low in the present invention.
3, the present invention develop water soluble drug nanometer Brain targeting preparation strengthen water soluble drug brain targeting, heighten the effect of a treatment, and improve its bioavailability.
The foregoing is only preferred embodiment of the present invention, not in order to limit the present invention, all any amendments done within the spirit and principles in the present invention, equivalent replacement and improvement etc., all should be included within protection scope of the present invention.

Claims (10)

1. the load of Brain targeting water soluble drug, is characterized in that, comprises pharmaceutical carrier propine acylation chitosan PA-CS-CHO, nitrine probe N-(2-(2-nitrine ethyoxyl) ethyl) palmitamide NAEP and LDL; LDL surface-assembled nitrine probe NAEP, the alkynyl on PA-CS-CHO and N 3nitrine on-LDL reacts and forms the amphipathic molecule of LDL targeting end as hydrophobic side.
2. Brain targeting water soluble drug load as claimed in claim 1, it is characterized in that, the structural formula of described propine acylation chitosan PA-CS-CHO is as follows:
Wherein, the scope of x is 2 ~ 30; The scope of y ' is 2 ~ 20; The scope of z is 8 ~ 30.
3. the preparation method of the Brain targeting water soluble drug load described in claim 1 or 2, is characterized in that, comprise the following steps:
By N-(2-(2-nitrine ethyoxyl) ethyl) palmitamide NAEP and low density lipoprotein, LDL LDL mixing in broad dose altogether, obtaining by self-assembling reaction the LDL that surface has nitrine probe, being labeled as N 3-LDL; Wherein, the mol ratio of described N-(2-(2-nitrine ethyoxyl) ethyl) palmitamide NAEP, low density lipoprotein, LDL LDL is 1: (0.5 ~ 2);
React under mild conditions by click chemistry, the alkynyl on propine acylation chitosan PA-CS-CHO and N 3nitrine on-LDL reacts, and obtains the load of Brain targeting water soluble drug; Wherein, described propine acylation chitosan PA-CS-CHO and N 3the mol ratio of-LDL is 1: (0.1 ~ 2).
4. the preparation method of Brain targeting water soluble drug load as claimed in claim 3, it is characterized in that, the preparation of described N-(2-(2-nitrine ethyoxyl) ethyl) palmitamide NAEP comprises the following steps:
(1) C 15h 31cONHCH 2cH 2oCH 2cH 2the synthesis of OH (NHEP)
In a nitrogen atmosphere, dimethyl formamide stirring and dissolving is added in Palmic acid, 2-(2-amino ethoxy) ethanol and triethylamine mixed liquor, after ice bath cooling, add the BTA-N of mixing in advance, N, N ', N '-tetramethylurea hexafluorophosphate and N-hydroxy benzo triazole, room temperature reaction, thin layer chromatography determination reaction end, obtains reactant solution; Wherein, described Palmic acid, 2-(2-amino ethoxy) ethanol, triethylamine, dimethyl formamide, BTA-N, N, the mol ratio of N ', N '-tetramethylurea hexafluorophosphate and N-hydroxy benzo triazole is 1: (1 ~ 1.5): (1 ~ 3): (1 ~ 5): (1 ~ 4.5): (1 ~ 6);
Reactant solution is dropwise instilled in the cold water in stirring, with chloroform extraction; The chloroformic solution collected uses citric acid solution successively, sodium bicarbonate solution, and saturated nacl aqueous solution washs, and revolve after anhydrous sodium sulfate drying and boil off chloroform and obtain crude product, crude product is recrystallization purifying in ethanol, vacuum drying, and exsiccator is preserved;
(2) C 15h 31cONHCH 2cH 2oCH 2cH 2the synthesis of OTs (PEMBS)
The NHEP that step (1) obtains is dissolved in chloroform, adds pyridine; After cooling, add p-methyl benzene sulfonic chloride, return back to room temperature, stirring is spent the night, and thin layer chromatography determination reaction end, obtains product; Wherein, the mol ratio of described NHEP, chloroform, pyridine, p-methyl benzene sulfonic chloride is 1: (1 ~ 6): (4 ~ 5.6): (1 ~ 1.4);
Product instilled in hydrochloric acid solution, centrifugal going out after supernatant obtains product; Product rinses with hydrochloric acid and water respectively, at room temperature vacuum drying, and exsiccator stores;
(3) C 15h 31cONHCH 2cH 2oCH 2cH 2n 3(NAEP) synthesis
The PEMBS obtained in step (2) is dissolved in dimethyl formamide under nitrogen protection, adds sodium azide, slowly heat, after backflow certain hour, be down to room temperature; Reaction mixture instills in frozen water in stirring, filters, and after water repeatedly flush cake, room temperature in vacuo is dry; Obtain end product by column chromatography purification, at room temperature vacuum drying, exsiccator stores; Wherein, the mol ratio of described PEMBS, dimethyl formamide, sodium azide is 1: (1 ~ 5): (4 ~ 5.6).
5. the preparation method of Brain targeting water soluble drug load as claimed in claim 3, it is characterized in that, the preparation of described propine acylation chitosan PA-CS-CHO comprises the following steps:
Under nitrogen protection, the NaAc_HAc buffer solution of chitosan and Potassium metaperiodate. are with stirring reaction at suitable ratio 4 DEG C; Then in solution, add appropriate ethylene glycol solution carry out stopped reaction; Product dialyses to remove impurity respectively after rotary evaporation is concentrated in NaCl aqueous solution and deionized water, and end product lyophilization, obtains hydroformylation chitosan CS-CHO; Wherein, the molal volume ratio between described chitosan, NaAc_HAc buffer solution, Potassium metaperiodate., ethylene glycol is 1: (1 ~ 5): (0.1 ~ 1): (0.1 ~ 1);
Under nitrogen protection, in acetylenecarboxylic acid aqueous solution, add N-hydroxy-succinamide, dropwise instill in hydroformylation chitosan CS-CHO aqueous solution after dissolving, after stirring, add 1-ethyl-(3-dimethylaminopropyl) carbodiimide hydrochloride; The mol ratio of hydroformylation chitosan, acetylenecarboxylic acid, N-hydroxy-succinamide and 1-ethyl-(3-dimethylaminopropyl) carbodiimide hydrochloride is 1: (1 ~ 4): (1 ~ 3): after (1 ~ 3) mixture reaction 10 ~ 72h, be transferred in bag filter, the unreacted micromolecule of dialysis removing in NaCl aqueous solution and deionized water, lyophilization obtains product.
6. a Brain targeting aqueous solubility pharmaceutical formulations, is characterized in that, comprises the Brain targeting water soluble drug load described in the claims 1 or 2 and Brain targeting water soluble drug; Wherein, the exposed Brain targeting aqueous solubility pharmaceutical formulations outside of LDL targeting end is formed as the amphipathic molecule of hydrophobic side and water soluble drug self assembly in the load of Brain targeting water soluble drug.
7. Brain targeting aqueous solubility pharmaceutical formulations as claimed in claim 6, it is characterized in that, the form of described Brain targeting aqueous solubility pharmaceutical formulations is the Nano microsphere of 35 ~ 100 nanometers;
Described Brain targeting water soluble drug is the diagnosis of central nervous system disease, macromole, the water-soluble substances for the treatment of needs, comprises lycoramine, galantamine, lycorine.
8. the preparation method of the Brain targeting aqueous solubility pharmaceutical formulations described in claim 6 or 7, it is characterized in that, comprise the following steps: the load of Brain targeting water soluble drug is dissolved in organic solvent and prepares organic solution, then the aqueous solution of described organic solution and Brain targeting water soluble drug is carried out self assembly in bag filter; Wherein, the volume ratio of the load of Brain targeting water soluble drug and organic solvent is 1: (1 ~ 10), and the WMCO=1000 of bag filter, described organic solvent comprises dimethyl sulfoxide, DMF, acetone, oxolane.
9. the application in Brain targeting aqueous solubility pharmaceutical formulations is being prepared in the Brain targeting water soluble drug load described in claim 1 or 2.
10. the application in central nervous system disease medicine is being prepared in the Brain targeting water soluble drug load described in claim 1 or 2.
CN201410423851.7A 2014-08-27 2014-08-27 Brain-targeted water soluble drug carrier as well as preparation method and application thereof Withdrawn CN104208709A (en)

Priority Applications (1)

Application Number Priority Date Filing Date Title
CN201410423851.7A CN104208709A (en) 2014-08-27 2014-08-27 Brain-targeted water soluble drug carrier as well as preparation method and application thereof

Applications Claiming Priority (1)

Application Number Priority Date Filing Date Title
CN201410423851.7A CN104208709A (en) 2014-08-27 2014-08-27 Brain-targeted water soluble drug carrier as well as preparation method and application thereof

Publications (1)

Publication Number Publication Date
CN104208709A true CN104208709A (en) 2014-12-17

Family

ID=52090880

Family Applications (1)

Application Number Title Priority Date Filing Date
CN201410423851.7A Withdrawn CN104208709A (en) 2014-08-27 2014-08-27 Brain-targeted water soluble drug carrier as well as preparation method and application thereof

Country Status (1)

Country Link
CN (1) CN104208709A (en)

Cited By (3)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN104587486A (en) * 2014-12-20 2015-05-06 盐城工学院 Chitosan-platinum (IV) prodrug conjugate and preparation method thereof
CN105853392A (en) * 2016-01-20 2016-08-17 深圳市老年医学研究所 Amaryllidaceous alkaloid targeted sustained-release preparation and preparation method and use thereof
CN110938156A (en) * 2019-12-16 2020-03-31 中国热带农业科学院农产品加工研究所 Amphiphilic chitosan, preparation method thereof and amphiphilic chitosan-based nano microcapsule applying amphiphilic chitosan

Cited By (4)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN104587486A (en) * 2014-12-20 2015-05-06 盐城工学院 Chitosan-platinum (IV) prodrug conjugate and preparation method thereof
CN105853392A (en) * 2016-01-20 2016-08-17 深圳市老年医学研究所 Amaryllidaceous alkaloid targeted sustained-release preparation and preparation method and use thereof
CN110938156A (en) * 2019-12-16 2020-03-31 中国热带农业科学院农产品加工研究所 Amphiphilic chitosan, preparation method thereof and amphiphilic chitosan-based nano microcapsule applying amphiphilic chitosan
CN110938156B (en) * 2019-12-16 2021-11-19 中国热带农业科学院农产品加工研究所 Amphiphilic chitosan, preparation method thereof and amphiphilic chitosan-based nano microcapsule applying amphiphilic chitosan

Similar Documents

Publication Publication Date Title
Liao et al. Dual pH-responsive-charge-reversal micelle platform for enhanced anticancer therapy
CN102988999B (en) Curcumin-polysaccharide conjugate as well as preparation method and application thereof
CN101791411B (en) Preparation and application of amphiphilic polysaccharide conjugate and pharmaceutical composition thereof
Hou et al. Smart nanocomposite hydrogels based on azo crosslinked graphene oxide for oral colon-specific drug delivery
Wang et al. Synthesis of an acid-labile polymeric prodrug DOX-acetal-PEG-acetal-DOX with high drug loading content for pH-triggered intracellular drug release
CN102060991B (en) Amphiphilic prodrug of 7- ethyl-10-hydroxycamptothecin and preparation method thereof
CN111848658B (en) Mitochondria-targeted fluoroborodipyrrole compound, preparation method and application of liposome-coated nano particles thereof
CN107417752A (en) One kind has compound of active anticancer and its preparation method and application
Chen et al. N-acetylneuraminic acid and chondroitin sulfate modified nanomicelles with ROS-sensitive H2S donor via targeting E-selectin receptor and CD44 receptor for the efficient therapy of atherosclerosis
CN104208709A (en) Brain-targeted water soluble drug carrier as well as preparation method and application thereof
CN113018450A (en) Drug carrier with tumor cell and tumor-related fibroblast double-targeting function, preparation method and application
Sun et al. Supramolecular nanomedicine for selective cancer therapy via sequential responsiveness to reactive oxygen species and glutathione
CN114796522A (en) Novel anti-tumor nano-drug for amplifying oxidative stress of targeted mitochondria
Liang et al. An Acid-Sensitive Nanofiber Conjugate Based on a Short Aromatic Peptide for Targeted Delivery of Doxorubicin in Liver Cancer
CN113577299B (en) ROS-responsive monoclonal antibody medicine oral nanoparticle and preparation method thereof
CN103933016B (en) A kind of capsaicin ternary nano micelle and method for making thereof and purposes
CN104045823B (en) A kind of Enoxolone derivative and its preparation method and application
CN101942098B (en) Amphiphilic chitosan with chemical crosslinking characteristic and preparation method thereof
JP7046321B2 (en) Modified styrene-maleic anhydride copolymer and its use
CN103055321A (en) Methoxy polyethylene glycol-polyphosphate diblock copolymer and adriamycin bonding medicine thereof
Hua et al. ROS-responsive nanoparticle delivery of ferroptosis inhibitor prodrug to facilitate mesenchymal stem cell-mediated spinal cord injury repair
CN112390872B (en) Keratin peptide derivative and preparation method, application and pharmaceutical composition thereof
CN112661961B (en) Amphiphilic polyoxazoline copolymer, and preparation method and application thereof
CN115192524B (en) Polymeric micelle encapsulating chain insoluble drug and preparation method and application thereof
CN102977358B (en) Module compound with hydrogen bond sequence specificity combination and preparation method thereof

Legal Events

Date Code Title Description
C06 Publication
PB01 Publication
SE01 Entry into force of request for substantive examination
SE01 Entry into force of request for substantive examination
WW01 Invention patent application withdrawn after publication
WW01 Invention patent application withdrawn after publication

Application publication date: 20141217